Ahlers I¹, Ahlersová E¹ , Bojková B¹ , Kubatka P¹ , Alberty R² , Adámeková E¹ , Marková M¹

Nimesulide (NIM) represents the non steroidal antiinflammatory drug, inhibitor of prostanoid synthesis, with marked influence on cyclooxygenase-2 activity. The aim of recent study was to determine the metabolic effect of this compound in rats, in the nycthemeral arrangement of the experiment. Male Wistar: han rats were kept under standard conditions, with light : dark regimen 12 : 12 h (light on at 7 a.m.). Nimesulide (Helsinn Healthcare SA, Lugan, Switzerland) was administered 1 hour after light onset (i.e. at 8 a.m.), or 1 hour after onset of darkness (i.e. at 8 p.m.) in the dose of 10 mg/kg b.w. subcutaneously. The control group was injected by the same volume of the vehicle, used for NIM dissolution (i.e. at 11 a.m. and 11 p.m.) the rats were sacrified and selected parameters of lipid and carbohydrate metabolism in the serum, liver, bone marrow and heart muscle were determined.

A marked increase in the serum glucose concentration, a moderate increase in serum cholesterol, liver triacylglycerol and bone marrow phospholipid concentration and a decrease in lipid peroxide concentration in the bone marrow was observed after NIM administration in the darkness. A decrease in liver glycogen and an increase in liver cholesterol and triacylglycerol concentration occured after NIM administration in the ligth part of day. Chronopharmacological studies for the determination of NIM effects are required in human subjects too.

Ahlersová E, Kassayová M , _majda B , Ahlers I

In the laboratory rat - exhibiting low photoperiodic activity - moderate response to various photoperiods on circadian oscillations of thyroid hormones were noted. In the present study we examined the effect of short artificial photoperiod LD 6 : 18 applied in a single season, on circadian variations of serum thyrotropin (TSH), thyroxine (T₄), triiodothyronine (T₃) and reverse T₃ (rT₃). The control group was kept in LD 12 : 12 regimen.

Male Wistar rats were adapted for five weeks to artificial light regimens LD 6 : 18 and 12 : 12. Cold light was switched on at 07.00 h in both regimens. The rats were examined at 3-hour intervals within 24 h, the serum concentration of THS, T₄, T₃ and rT₃ were determined radioimmunochemically. The results were evaluated by the cosinor analysis with a chosen 24 h and 12 h rhythm periods. In the short day the circadian oscillations of TSH, T₄ and rT₃ did not show rhythmicity except of T₃ with rhythm detection. Circadian oscillations of TSH and thyroid hormones examined in LD 12 : 12 regimen were rhythmic, T₄ rhythm was of borderline significance. The mesor values of TSH, T₄, T₃ resp. rT₃ were significantly higher in the LD 6 : 18 regimen in comparison with values of control group (LD 12 : 12 regimen).

The effect of short artificial photoperiod applied in spring influenced circadian oscillations of serum TSH and thyroid hormones: the mesors were remarkably higher in comparison with LD 12 : 12 regimen values, the rhythmicity was not present with the exception of T₃.

Babinská K¹, Klvanová J², Béderová A³

The fatty acid intake and plasma fatty acid composition were assessed in a group of iron deficient subjects (n=25, mean age 42,8±8,3y) and in a control group of matched healthy subjects (n=25, mean age 44,5±12y). Serum iron levels were assessed spectrophotometrically, plasma fatty acids were determined by gas chromatography. Fatty acid consumption was estimated from 24-hour dietary recalls.

The intake of essential fatty acids (18:2n6 and 18:3n3) did not differ significantly between both groups. Intake of 18:3n3 (1,5 g per day) was lower than the recommendation. Similarly, extremely low consumption of LCPUFA n-3 (20:5n3 and 22:6n3) was observed. Lower (p<0,04) fatty acid index (calculated as product to substrate ratio) for the formation of g-linolenic acid (18:3n6/18:2n6) and dihomo-g-linolenic acid (20:3n6/18:2n6) was found in the iron deficient group. Serum iron level significantly correlated with the ratio 18:3n6/18:2n6 (r=0,33; p<0,03), as well with 20:3n6/18:2n6 (r=0,33; p<0,03). Significantly lower plasma level of 20:5n3 was observed in the iron deficient group.

Bahníková M, Matějovič P, Pásek M, _imurdová M, _imurda J

Ajmaline, a derivative of rauwolfia, is known as a potent antiarrhythmic drug currently used in the treatment of a variety of both ventricular and atrial tachyarrhythmias. It is regarded as a representative of class Ia according to Vaughan Williams classification. Numerous clinical studies have shown that ajmalin slows conduction of excitation, prolongs the refractory period, QT and QRS interval. Surprisingly, only sporadic cellular electrophysiological data are available.

The whole cell patch clamp technique was applied to rat ventricular myocytes at room temperature. Ajmaline in the range 3.10^-7 - 5.10^-3 M was found to suppress three currents: the fast sodium current (I_Na), 4-aminopyridine sensitive transient outward current (I_to) and current measured at the end of the imposed 0.1 s rectangular pulses (I_K) in a dose-dependent manner with EC₅₀ of 8.2, 27.5, and 166 mM and the Hill coefficient of 0.62, 0.67, and 0.44, respectively. Although the block of ajmaline on I_Na and I_to was affected by depolarization considerably, it did not show significant frequency dependency (0.33 - 3.3 Hz). The effect was reversible in all concentrations applied. The half times of the onset of the block action was less than 150 s for all the explored current components. A significant voltage dependence of I_K block was not observed.

This study demonstrates low blocking specificity and frequency independent effect of ajmalin in rat ventricular myocytes. The latter corresponds well with the observed fast equilibration of block level following repolarization.

Barancik M^1,2, Strohm C², Bruehl v.ML², Schaper W^2 
1Inst Heart Res, Slovak Acad Sci, Bratislava, Slovak Republic, and ²Dept Experim Cardiol, Max-Planck-Instit, Bad Nauheim, Germany

Evidence accumulates suggesting that mitogen-activated protein kinases (MAPKs) could play an important role in the stress responses and for survival and cell death. We investigated the role of MAPK cascades in the ischemic preconditioning (IP) mediated cardioprotection in pig myocardium. Methods: The hearts (open chest model) were subjected to regional ischemia. IP was induced by two cycles of 10 min LAD-occlusion and 10 min reperfusion, prior to the onset of prolonged index ischemia. "Stress" kinase activator (anisomycin- AN), specific inhibitors (PD98059, UO126 for ERKs; SB203580 for p38-MAPK) and actinomycin-D (Act-D) were infused systemically or intramyocardially. The infarct size (IS) measurements, kinase and Western blot assays were performed as described previously (1). Results: IP significantly reduced the IS. During the IP procedure an activation of ERKs, JNKs and down-regulation of p38-MAPK activities was observed. Infusions of PD, UO and Act-D significantly reduced the IP-induced cardioprotection and this was connected with inhibition of ERK activities. Inhibition of p38-MAPK cascade by SB significantly reduced IS after index ischemia but did not influence the IS reduction mediated by IP. AN reduced IS when infused prior to index ischemia and this was associated with a specific activation of JNKs. Conclusion: The results point to the positive role of ERKs and SAPK/JNKs in adaptive responses of myocardium to ischemia. On the other hand, p38-MAPK activation is believed to accelerate the death pathway, the inhibition of which shifts the balance toward cell survival.

1. Barancik et al., J Cardiovasc Pharmacol 35; 2000:474-483

Barcal J, Jelínková D, _tenglová V, Vožeh F, Žalud V

Lurcher mutant mice as a model of cerebellar degeneration were used. Because our previous studies have shown important changes not only in the cognitive functions but also in the level of excitability, this work is focused on the hippocampal activity (electrically evoked, LTP). Healthy mice ("wild-type") were used as a control group. Hippocampal potentiation was performed in acute experiments under urethane anesthesia (20%, 2g/kg). For stimulation (perforant path: lambda—3.0-1,9) and registration (ipsilateral dentate gyrus 2.0 -1.7-2.0) stainless steel electrodes were used. Biphasic pulses 3V, basal low frequency 0.1 Hz, duration 0.1 ms, high-frequency stimulation (HFS) 100Hz, 10 bursts each 10 s were applied. On the contralateral side L-arginine (substrate for NO synthesis) and nitro-L-arginine (blockator of NO-synthase) were administered intracerebroventricularly (1ml, 30 min after HFS). In the selected animals NADPH-diaphorase positivity has been used as an NOS marker. The block of NOS impaired evoked responses in both groups of animals (wild and Lurcher) whereas increase of NO bioavailability (after L-arginine administration) caused an enhancement of responses, but only in healthy animals with statistically significant effect. Taken together, the cognitive deficit in this cerebellar degeneration is characterized by the change of NOS activity in hippocampal region. Results after influencing the nitric oxide bioavailability suggest that NO plays an important role (retrograde messenger?) in various types of the hippocampal activity in our cerebellar degeneration model.

Béder I¹, Kittová M¹, Maťa_eje A¹, Čársky J², Országhová Z², Babinská K ¹

Aminoguanidine (AG) and its adducts with aldehydes (Schiff bases) are membranal active substances.

The aim of this work was to observe the effect of AG, its adduct with pyridoxal-pyridoxylidenaminoguanidine (PAG) and pyridoxal on erythrocyte elasticity with expected differentiated protective influence. In 7 non-diabetic and 16 diabetic patients with insulin dependent diabetes mellitus erythrocyte elasticity was estimated by determination of their deformability using filtration method with centrifugation. Other hematological variables including erythrocyte and reticulocyte counts, hematocrit values, hemoglobin concentration and the mean cell volume were determined. AG improved erythrocyte filterability by 4%, aminoguanidine derivate PAG by 11% and pyridoxal by 13% in healthy subjects. In diabetic patients the AG effect on erythrocyte filtrability was improved by 7%, PAG effect by 9 % and pyridoxal effect by 15% in comparison to the control group. The other investigated hematological variables in both groups were within the range of the physiological standard.

Beňačka R ¹, Tomori Z ¹

Introduction: The present work was designed to compare the resuscitation effects of mechanical, air-puff and/or electrical stimulation of epipharyngeal (EPS), inferior nasal (INS) & nasal philtre areas (NPS) during respiro-circulatory failure induced by peracute systemic anoxia, asphyxia and/or short brain ischaemia.

Methods: In 14 anaesthetised nonparalyzed cats we analysed diaphragmal EMG, heart rate (HR), systemic blood pressure (BP), native & spectral EEGs, brainstem auditory/ median or trigeminal sensory evoked potentials (EP), trachel air-flows and ABS.

Results: EPS could revive respiration within 90s of apnoea Recovery was indicated by instantaneous series of aspiration reflexes (ARs) (316±12 ms, M±SE). This was followed by immediate rise in both BP (2-3x) and HR (4x) and sudden restore of pontine EP. EEG revived within next 20-40s. Rhythmic breathing reappeared 40-60 s after EPS, while thalamocortical EP recovered with 80-120s delay. Bilateral carotic occlusion (BCO) abolished renewal of brain activity in spite of present AR and sustained BP. INS evoked similar effect to EPS. although less effective (only in 63% at 30-60 s of apnoea). NPS could reactivate cardiovascular, respiratory and brain responses even 120-150 s after apnoea onset Recovery started with series of 4-10 AR-like bursts (216±12ms) followed by 2-4x increase in HR and BP and periodic gasps resuming into normal rhythm. When BP>30-40 mmHg repetitive NPS could preserve automatic breathing even after cortical death induced by BCO.

Benick_ J¹, Najvirtová M¹, Baqi L¹, Križanová O², _trbák V¹

Introduction: Neonatal streptozotocin (nSTZ) treatment in rats leads to parallel depletion of pancreatic insulin and TRH followed by spontaneous regeneration of insulin- but not TRH-containing cells. An impaired insulin response to glucose stimulation persists in nSTZ rats for the rest of life, which is the similar effect as that observed in mice with prepro-TRH gene knock-out. We therefore used the model of nSTZ rats to determine the possible role of perinatal absence of TRH in the development of insulin responsiveness dysfunction.

Methods: At the day of birth the rats were injected with 90 mg/g BW STZ i.p. TRH (10 ng/g BW/day s.c.) was injected daily during the first week of life and function of isolated pancreatic islets was studied at the age 12-14 weeks. Plasma glucose, content of insulin in plasma and in pancreatic islets, and insulin release in vitro were measured by radioimmunoassay.

Results: The nSTZ rats had normal glycemia and insulinemia. Insulin content in islets was lowered and we observed no insulin response to glucose and high KCl stimulation in vitro in nSTZ animals. Perinatal TRH treatment deeply decreased protein and insulin content in islets of both control and nSTZ rats, and enhanced basal insulin secretion in nSTZ animals in vitro. Insulin response to glucose stimulation reappeared only in TRH-treated nSTZ females.

Beręsewicz A, Mączewski M, Duda M

Cardiac ischemia/reperfusion (IR) causes damage to cardiomyocytes and coronary endothelium, the changes preventable by ischemic preconditioning (IPC). Adverse consequences of endothelial dysfunction (ED) include increased granulocyte accumulation within IR myocardium. In this study the hypothesis was tested that IPC and diazoxide, an activator of mitochondrial K_ATP channel, confer protection against ED via the mechanism that involves attenuation of myocardial peroxynitrite formation (a product of the reaction: superoxide (O₂^-) + NO). This may involve decreased O₂^- production, increased O₂^- removal, and/or decreased NO production. Perfused guinea-pig hearts were subjected either to (i) 30 min/30 min IR or to (ii) 3x5 min IPC prior to IR; (iii) 3x5 min diazoxide infusion/washout prior to IR or (iv) the hearts were infused for 10 min before the ischemia and during the initial 10 min of the reperfusion with either SOD or L-NMMA. Coronary flow responses to acetylcholine served as measures of endothelial function. IR impaired acetylocholine response by 62%, an effect prevented by IPC, diazoxide, SOD, and L-NMMA. The reperfusion-induced outflow of O₂^- was reduced, that of .OH increased, and that of NO not affected by IPC, diazoxide and SOD. Thus, it is enhanced O₂^- removal rather then decreased O₂^- production that accounts for the reduction in O₂^- outflow in IPC and diazoxide groups (perhaps via increased activation of mitochondrial K_ATP and then activation of SOD). Reperfusion-induced outflow of NO was reduced and that of O₂^- and .OH not affected by L-NMMA. These results implicate that it was the product of the reaction O₂^- + NO, but not O₂^-, per se, that mediated ED in IR guinea-pig heart, and that IPC and diazoxide afforded endothelial protection because they attenuated the biological availability of O₂^-, and thereby of peroxynitrite.

Biernat J, Sendur R, Pawlik M, Obuchowicz R, Zejc-Bajsarowicz M, Pawlik WW

The aim of our study was to evaluate the role of sensory neurons and sensory peptides in the protective action against CCl₄ induced liver injury.

Experiments were performed on anesthetized rats, weighing 220-240 g. Hepatic blood flow (HBF) was determined by Laser Doppler flowmetry and arterial pressure (AP) was measured. AspAT and AlAT serum levels were measured 24 hours after CCl₄ 

The first group: placebo, in the second sensory denervation was made by administration of Capsaicin (CAP) in increasing doses. In the third group carbon tetrachloride (CCl₄) alone (0,3 ml/kg s.c.) was given. In the fourth group CCl₄ was given 14 days after capsaicin (CAP) s.c. In the fifth and the sixth group CCl₄ was administered after CGRP (0,16 mg/kg i.p.) and Substance P (0,1mg/kg i.p.)respectively.

CCl₄ (0,3 ml/kg s.c.) produced liver damage in rats which was manifested at 24 hr in serum rise of AspAT and ALAT to 485±120 U/L and 368±83 U/L respectively. AP and HBF were decreased by 15±7% and 25±6% respectively. In chronically CAP denervated rats: CCl₄ produced significantly less hepatic damage as occurred in a rise of AspAT and ALAT to 358±45 and 255±38 U/L respectively, while HBF was increased by 58±8%. Pretreatment with CGRP and substance P did not significantly changed AspAT¢s and ALAT¢s values but HBF increased by 19±4% and 24±6% respectively in comparison to CCl₄ alone.

Biliczki P¹, L. Virág¹, N. Iost², Papp JGy^1,2, Varró A¹:

The aim of this study was to investigate the possible role of the repolarization reserve and the interaction of different potassium channels in the dog ventricular muscle preparations, by applying the conventional microelectrode and whole cell patch-clamp techniques at 37 °C. Complete block of I_Kr by 1 µM dofetilide (DOF) lengthened action potential duration (APD) by 45.6±3.6 %, n=13 (at cycle length CL=5000 ms). Chromanol 293B (CRO) at 10 µM (concentration which selectively blocks I_Ks current) applied alone did not markedly lengthen APD (< 7 %), but when repolarization was already prolonged by complete I_Kr block (1 µM DOF), inhibition of I_Ks with 10 µM CRO substantially delayed repolarization (38.5±8.2 %, n=6, at CL=5000 ms). BaCl₂ (BA) applied alone in the concentration of 10 µM (which selectively blocks I_k1 current) lengthened APD by 33.0±3.1 % (n=11), but when I_Kr was blocked (1 µM DOF) 10 µM BA produced an excessive frequency dependent lengthening in APD by 104.3±22.1 % (n=7), frequently initiating early afterdepolarizations (EAD). We have concluded that in the dog ventricular muscle when only one type of potassium channels is inhibited, excessive APD lengthening is not likely to occur. Dog ventricular myocytes seem to repolarize with a strong safety margin ("repolarization reserve"). However, when this normal "repolarization reserve" is attenuated (due to drugs, remodelling or genetic disorders), the otherwise minimal or moderate potassium current inhibition can result in excessive and potentially proarrhythmic prolongation of the ventricular action potential duration. Therefore, application of drugs which are able to block more than one type of potassium channels is probably more hazardous than the use of specific inhibitors of a given potassium channel.

CIRCULATORY, RESPIRATORY AND OTHER MARKERS AT HEMORRHAGIC HYPOVOLEMIA.

Bračoková I, _vorc P, Dorko E, Kassayová K, _timmelová J

Breier A¹, Barančík M², Kvačkajová J², Boháčová V²

Mouse leukemic cell line L1210/VCR used in our studies were prepared by adaptation of parental sensitive L1210 cell line to vincristine. In L1210/VCR cells was observed also increased cross-resistance to other cytostatics such as vinblastine, doxorubicin, mitomycin C, and actinomycin D. Important factors influencing the realization of MDR in L1210/VCR cells are flexibility of structure, lipophilicity and molecular weight of used cytostatics. Resistance to vincristine and doxorubicin was found to be associated with decreased intracellular accumulation of these drugs. Multidrug resistant cell line L1210/VCR is characterized by overexpression of PGP (determined at protein and mRNA levels) but not by an increase of activities of glutathione S-transferase. Moreover, the exposure of resistant cells to several cytostatics (vincristine, doxorubicin, mitomycin C) did not influence the activities of GST. Several drugs (chemosensitisers, like: calcium entry blockers, inhibitors of calmodulin, local anesthetics and some xantine derivatives) were found to depress the P-glycoprotein mediated MDR. The primary structure of PGP contains sequences with consensus for phosphorylation sites of some protein kinases and the linker region of MDR1-PGP was found to be phosphorylated in vivo within the acidic domain. Some protein kinase activators (phorbol myristate: PKC) and inhibitors (bisindolylmaleimid: PKC; SB203580: p38-MAPK; PD098059: ERKs) on the resistance of L1210/VCR cells were found to influence the MDR of our cells. Development of MDR in L1210/VCR cell line was associated with significant changes in content, phosphorylation and activity of p38-MAPK. These facts indicate that the phosphorylation of PGP represents important mechanism involved in the regulation of its transport activity.

Brezová A¹, Uhrík B¹, Heizmann CW²

S100A1 protein is a small acidic protein with two EF-hand type Ca²⁺-binding motifs. S100A1 can form homodimers (slow skeletal and heart muscles, kidney) or heterodimers with S100B (brain). Function and localization of S100A1 has not been clearly revealed yet, but it is assumed to have regulatory effects along the Ca²⁺-signal transmission pathway. The aim of our work was to study the subcellular localization of S100A1 protein in normal rat heart.

Rat hearts were fixed by coronary perfusion with low aldehyde concentration solution. Samples from left and right ventricles and left and right atriums were dissected and processed according to Tokuyasu technique. Ultrathin cryo-sections were immunolabelled and examined in fluorescent and electron microscope. Immunolabelling was performed with polyclonal rabbit IgG against human recombinant S100A1 diluted 1:100. The specifity of primary antibody was tested by saturated primary antibody and by rabbit normal serum.

In sections from all types of heart tissue the most pronounced immunolabelling was seen in mitochondria. Much lower labelling could be detected in myofibrils, at different levels of sarcomere. Up to the present no target proteins for S100A1 are known in mitochondria. However, the functional significance of S100A1 for cardiac muscle may be assumed from results of studies on patologically changed heart: a lowered expression of S100A1 in human cardiomyopathy and its right ventricular upregulation in chronic pulmonary hypertension.

Brtko J¹, Podoba J², Schmutzler C³, Köhrle J³

The majority of thyroid adenomas are of clonal origin. The present study was undertaken to investigate the status of functional thyroid hormone receptors, major thyroid hormone signal mediators, in both the human TAs and CNs in comparison with a normal thyroid tissue from the same patient. Electrophoretic mobility shift assays using a DR4 („direct repeats" 4), a thyroid hormone responsive element (TRE) derived from the human type I iodothyronine 5´-deiodinase promoter demonstrated the DNA-binding of thyroid hormone receptors (TRs) in thyroid tissue nuclear extracts. A significant increase (p ‹ 0.05) in the functional binding properties of TRs to the DR4 TRE was found in TAs when compared to normal thyroid tissue. Contrary, a marked diminution in the TR-TRE complex formation was found in CNs in comparison with normal thyroid tissue. In addition, functional activity of the iodothyronine 5´-deiodinase (5´-DI) was analyzed in benign tumours, thyroid TAs and CNs in comparison with that of normal thyroid tissue. A significantly increased (p ‹ 0.01) activity of 5´-DI was demonstrated in TAs, and in contrast, decreased values of the enzyme activity were found in CNs when compared to a normal tissue.

From the data it is suggested that both the status of TR-TRE complex formation and the activity of the 5´-DI may be altered in benign tumours of human thyroid gland.

Cagalinec M^1,3, Matea_ik A³, Chorvátová A^2,3, Chorvát D³

In recent decades we could see evident progress in biological optical microscopy methods, caused by wide availability of lasers, CCD detectors and digital data processing. Meanwhile, there is an apparent paradigm shift from applications using simple measurement of light intensity to the utilisation of derived, more complex fluorescence parameters like excited state lifetime, anisotropy etc.

Here, we present an extension of mentioned philosophy towards characterisation of spatially resolved calcium waves in isolated left ventricular rat cardiomyocytes, labelled with fluorescence calcium indicator Fluo-3. In addition to CCD imaging providing series of fluorescence images, we have constructed normalised temporal gradient of fluorescence intensity in each pixel and find it’s extremes describing local dynamic characteristics of the calcium wave (rise and decay constant). Using this methodology it is possible to obtain spatially resolved images of cell function parameters instead of simple calcium concentration mapping. We describe a simplified kinetic model that is possible to use for analysis of fluorescence data practically in real time, what is essential regarding possible future use of this setup in regular experiments for cellular physiology or cellular pharmacology.

The translocation of GLUT4 glucose transporter protein to the plasma membrane (PM) 30 min after oral application of glucose (3g/kg b.wt.) after overnight fasting was studied in insulin resistant hereditary hypertriglyceridemic rats (HHTg) in epidydimal fat pad, diaphragm, m. gastrocnemius and heart. In addition, the glycogen content in muscles was examined as one of the indicators of glucose utilisation. The adult HHTg males and control (C) males were kept one week on high sucrose diet. After the diet the HHTg group had worse glucose tolerance compared with control group and significantly higher content of serum triglycerides (3,7 ± 0,1 vs 1,6 ± 0,2 mmol/l).

The major differences in the GLUT4 content in PM between HHTg and control groups after glucose load were found in adipose tissue (the increase of 29% vs 90%). In m. gastrocnemius of control group the GLUT4 content increased by 50%, in diaphragm by 37% whereas in HHTg group we found no changes. GLUT4 content in myocardium was not different between HHTg and control rats. The changes in glycogen content in m. gastrocnemius reflected the changes in PM GLUT4 content.

We conclude that the impaired sensitivity of tissues to insulin in HHTg rats is associated with impaired ability to translocate GLUT4 transporters to the plasma membrane and the decreased synthesis of glycogen in skeletal muscle.

Pulmonary surfactant is a film of surface active agents that coats the alveolar surface and the small conducting airways. The physiological advantages of lung surfactant are that (i) it reduces the work of breathing and thereby reduces the muscular effort needed to expand the lungs; (ii) it lowers the surface tension at low lung volume, thus preventing the alveoli to collapse at the end of expiration; (iii) it prevents the deflation of individual alveoli at different rates.

Natural surfactant is a mixture of lipids, proteins (about 10%), and a small portion (less than 1%) of carbohydrates. The major lipid component is dipalmitoylphosphatidylcholine (DPPC). Adequate surface activity in vivo is only found in surfactant preparations that contain phospholipids and specific, surfactant associated proteins (SP). Four surfactant proteins have been described to date (SP-A, SP-B, SP-C, SP-D).

Surfactant is synthesised, stored, and recycled or catabolized by alveolar type II epithelial cells. It is packed in dense lamellar bodies, after exocytosis converted to tubular myelin in alveolar space and spread into a film at air/liquid interface. Control of metabolism of surfactant takes part at local, humoral and neural levels.

Camborova P¹,Pechan J², Ferak I³, Hubka P⁴, Jakubovsky J⁵, Hulin I¹.

The relation between gastric electrical activity measured at the surface of the skin and contractile activity is one of the complications of electrogastrography (EGG). Dominant frequency of the gastric slow wave in the EGG determins the maximum frequency of the contractions and could reflect the activity of the ICC network. The aim of our study was to detect the changes in stomach electrical activity in a patient with gastric outlet obstruction lasting 2 years. The EGG recordings were performed before the surgical correction and 10 monthsafter the operation.Before the surgical correction stable electrical activity was recorded (2.5cpm) and ultrasound revealed absence of gastric contractions. This electrical activity was slightly lower in comparison with mean values in our group of healthy subjects. 10 months following the surgery activity of 2 cpm was observed..The surgery confirmed gastric dilatation with major curvature length of 80 cm. Histology revealed mild dysplasia, chronic gastritis and Helicobacter pylori.

Even enormous gastric dilatation and structural changes haven’t resulted in a loss of electrical activity. The question is, why has the synchronized electrical activity not initiated gastric contractions.

Cendelín J¹, Žalud V¹, Jelínková D¹, _tenglová V¹, Vožeh F¹, Vrba J²

We studied the effect of high frequency electromagnetic field (HF EMF) on healthy wild type (+/+) and Lurcher mutant (+/Lc) mice (C3H strain). Lurcher mutants served as a model of olivocerebellar degeneration.

Spatial learning ability of adult animals was tested immediately after acute exposure to HF EMF or control conditions. Young mice were exposed to long-term influence (postnatal days 2 — 21) of HF EMF that involved the rapid brain growth spurt. Spatial learning was tested using Morris water maze. Brains of selected animals were processed histochemically and immunohistochemically to detect NADPH diaphorase and c-Fos.

Both +/+ and +/Lc adult animals exhibited no changes of spatial learning ability after acute exposure to HF EMF. The long-term exposure caused deterioration of spatial learning ability in +/+ and it slightly improved results in +/Lc. These differences were statistically insignificant. Detection of NADPH diaphorase showed no differences between exposed and control adult mice. In young control +/Lc was higher NADPH diaphorase activity in hippocampus and in some other brain structures as compared with exposed mice. Certain changes of c-Fos immunoreactivity in dependence on HF EMF exposition also were found.

We found no significant effect of HF EMF. However, some changes were discovered in mice exposed to HF EMF during their early ontogenetic development.

Cernayova A, Pancza D, Ravingerova T

Myocardial adaptation to ischemia has been recently shown to trigger multiple signaling pathways including activation of receptor tyrosine kinase and subsequent stimulation of p38-MAP kinase pathway. The aims of the study were to investigate their role in the protective effect of ischemic preconditioning (IP).

Isolated Langendorff-perfused rat hearts were subjected to test ischemic challenge (TI) induced by 25 min global ischemia (GI) followed by 35 min reperfusion (R). Recovery of function at the end of reperfusion (left ventricular developed pressure, LVDP) expressed as % of the initial value served as the end-point of injury. The hearts were adapted to ischemia by 2 episodes of 5 min GI and 5 min R before TI. To block receptor tyrosine kinase pathway, the hearts were pretreated with genistein (50 and 100 µM), and SB 203580 (5 µM) was used to inhibit p38-MAP kinase. Both drugs were applied 10 min before the onset of ischemia.

After TI/R, LVDP in the control group was 22.6 ± 3.5%, whereas preconditioned hearts showed a significantly better recovery of function (LVDP 63.0 ± 5.1%; P<0.05). 100 µM genistein partially blocked protective effect of IP, whereas SB 203580 completely abolished protection afforded by IP (LVDP 27.7 ± 7.8%).

IP protects rat hearts against myocardial dysfunction caused by ischemia/reperfusion. Blockade of cardioprotection with SB 203580 suggests a potential role of the activation of p38-MAP kinase pathway in the protective effect of IP. The activation of the upstream signaling mechanisms cannot be excluded.

Červenáková Ž¹, Penesová A¹, Ježová D¹, Kvetňansk_ R¹, Hinghofer-Szalkay H², Macho L¹, Ko_ka J¹

Changes in body fluid distribution are known to influence endocrine gland function. We studied the effects of altered plasma volume (PV), achieved by different body positions, on neuroendocrine response to insulin-induced hypoglycaemia.

Tests were performed in 12 subjects on 2 occasions: "head-up" (+60º head-up tilt standing for 30 min and hypoglycaemia in sitting position afterwards) and "leg-up" (leg-up position for 30 min and hypoglycaemia in leg-up position afterwards) in a random order. Controlled hypoglycaemia was induced by insulin infusion and adjusted to 2.7 mmol/l for 15 min by glucose infusion.

PV was significantly greater in leg-up (p<0.001) and lower in head-up (p<0.001) position than basal value. Head-up position caused increases in ACTH, aldosterone, noradrenaline (NA) and PRA (p<0.01). Leg-up position resulted in decreases in GH and adrenaline (A) (p<0.05). Contraregulatory response to controlled hypoglycaemia was mild. The significant increase in GH and A responses to hypoglycaemia used did not differ between the positions. Hypoglycaemia failed to activate ACTH release in head-up position. As to the gender differences, A response to hypoglycaemia was greater in males than in females (p<0.001).

&#These results show that reduced PV is associated with elevated plasma NA, ACTH, aldosterone and PRA while augmented PV with decreased A and GH concentrations. In the case of ACTH, the first stimulus is consequential for the subsequent response. Rise in A levels during hypoglycaemia depends on gender and is decreased in women.

Červinková Z, Radvaková D

&#The aim of our study was to evaluate impact of bile duct ligation on the functional and morphological parameters of small intestine.

Material and methods: The experiments were performed on 30 male albino Wistar rats with an initial body mass of 210-225 g. In the first group secondary biliary cirrhosis (BC) was induced by ligation of bile duct, the second control group (C) underwent median laparotomy. The rats were sacrificed three weeks after the operations. The extent of liver damage and liver regeneration was determined by assessment of serum activities of AST, ALT, GMP and AP, serum concentrations of albumin and CRP, liver DNA synthesis, and mitotic activity of hepatocytes. Intestine permeability was measured by lactulose-mannitol test, morphological changes were evaluated using histological estimation.

Results: Significant increase (p<0.05) of biochemical parameters together with liver morphological changes clearly documented development of BC in rats with bile duct ligation. Lactulose-mannitol test and intestine DNA synthesis were significantly increased (p<0.05) in BC rats to compare with controls.

Conclusion: Secondary biliary cirrhosis induced injury of small intestine shown by histological (confluence and irregularity of villi) and functional changes (impairment of intestinal barrier). These changes are probably to some extent related to the presence of portal hypertension. Intestine injury was followed by induction of reparative process judging by increased DNA synthesis in the small intestine.

Chvátal A, Anděrová M, Syková E

In conclusion, our experimental data and mathematical analysis reveal a more compact ECS volume around oligodendrocytes than around astrocytes. Such heterogeneities in the gray matter may selectively affect the diffusion of neuroactive substances in specific areas and directions and facilitate spatial K⁺ buffering in the nervous tissue.

Czakóová G¹, Kasalová L², Novotná J², Hermochová D¹, Korynta J³, Herget J¹

We have shown in the rat lungs that chronic hypoxia initiates increase in lung collagenolytic activity which results in the presence of low molecule weight collagen type I cleavages . The oxygenation of cornea is provided mainly directly from the athmosferic air. Therefore the minimal participation of the blood born substances may be anticipated in hypoxic corneal tissue. The aim of our study was to find the low molecular collagen peptides in the corneas of rats exposed to hypoxia. The rats were exposed to isobaric chronic hypoxia 10%O₂ for 4 and 14 days. Gel electrophoretic separations (SDS-PAGE) of the collagenous proteins isolated by limited pepsin digestion from corneas of 6 rats exposed to 4 days hypoxia, 6 rats exposed to 14 days hypoxia and 6 and 6 respective normoxic controls were performed. We found no differences in gel electrophoretic prophile of collagenous proteins between the normoxic rats and groups of rats exposed to hypoxia. There were no signs of vascularization of corneal tissue in rats exposed to hypoxia. We conclude that it is unlikely that the presence of collagenous cleavages in chronic hypoxia is a direct effect of lack of oxygen on collagenous matric proteins.

1.

Insuline-like growth factor-1 (IGF) overexpression was reported in acute pancreatitis. The aim of our studies was to determine the effect of IGF administration on the development of caerulein-induced pancreatitis (CIP).

Acute pancreatitis was induced by s.c. infusion of caerulein (10 mg/kg/h) for 5 h. IGF was administrated twice (30 min prior to caerulein or saline infusion and 3 h later) at the doses: 2, 10 or 50 mg/kg s.c. Immediately after cessation of caerulein or saline infusion the pancreatic blood flow (PBF), plasma amylase and lipase activity, plasma interleukin-1b (IL-1b) and interleukin 10 (IL-10) concentration, cell proliferation, and morphological signs of pancreatitis were examined.

Administration of IGF without induction of CIP increased plasma IL-10. Treatment with IGF, during induction of pancreatitis, increased plasma IL-10 and attenuated the pancreatic damage, what was manifested by histological improvement of pancreatic integrity, the partial reversion of the drop in DNA synthesis and PBF, and the reduction in pancreatitis evoked increase in plasma amylase, lipase and IL-1b level. Protective effect of IGF administration was dose-dependent, the dose 2 x 50 mg/kg caused the maximal reduction in pancreatic damage.

Conclusions: (1) Administration of IGF attenuates pancreatic damage in CIP; (2) This effect seems to be related to the increase in production of IL-10, reduction in release of IL-1β, the improvement of PBF and the stimulation of pancreatic tissue growth.

Donič V, Doničová V, Le_ko R, Tomori Z

Many sleep laboratories reported elevated blood pressure (BP) and changes in heart rate variability (HRV) of patients with sleep related breathing disorders (SRBD). These changes are not always clearly visible by using standard polysomnographic recordings. Sleep apnoea patients create usually not a homogenous group, but may have obstructive sleep apnoea syndrome (OSAS), OSAS treated with CPAP, upper airway resistance syndrome, simple snoring, etc. Therefore 24-h ambulatory (BP) devices, which offer blood pressure and HRV assessment during sleep, but also during wake period of the day, could be useful for study and better understanding of this phenomena.

Methods: Heart rate and BP were monitored using 24-h ambulatory blood pressure device (Cardiotens, Meditech Budapest) in adults and power spectral analysis of HRV was performed. Low frequency/high frequency ratio LF/HF, which reflects the sympatho-vagal balance, was calculated for the period of sleep vs. wakefulness (S/W). Our aim was 1) to detect the sensitivity - utility of our parameter in a group of patients with differentiation of OSAS against a group treated by CPAP, with respect to sympatho-vagal tone, 2) to select patients with high probability of SRBD

Results: Increased S/W ratio reflecting sympathetic activation was observed in patients with OSAS having (RDI 43.8) (0.9), compared to patients treated with CPAP (0.68), and people without SRBD.

Distribution of GABAergic neurons in the auditory cortex and in subcortical auditory structures.

Druga R, Ouda L, Syka J

Gamma-aminobutyric acid (GABA) is considered the major inhibitory transmitter in the CNS. The distribution of GABA-producing neurons in the auditory cortex (AC), the medial geniculate body (MGB) and the inferior colliculus (IC) of the rat was investigated by using an antibody to GAD and GABA in effort to obtain normative data for future experiments.

The IC contained a large population of GABA immunoreactive neurons distributed throughout all this subdivisions. A higher density of GABAergic neurons was evident in the basal part of the central nucleus and in the basal part of the external cortex of the IC.

The density of GABAergic neurons within the MGB was lower than in the IC and GABAergic neurons prevailed in the ventral division of the MGB.

In the auditory cortex GABAegic neurons were found in all fields (Te 1, Te 3 and Te 2 ) and were distributed in all cortical layers, particularly in layers IV and VI.

Dvorakova M^1,2, Kummer W¹

1Institute for Anatomy and Cell Biology, Justus-Liebig-University, Giessen, Germany, and

Eckhardt A^1,4, Mik_ík I^1,4, Cserháti T², Forgács E², Zicha J¹, Deyl Z^1,3

Depository effects in collagen (and slowly metabolized proteins in general), are very difficult to assess owing to extremely low concentration of the modifying (deposited) entities in the protein matrix. Typical of these nonenzymatic changes is glycation (Maillard reaction). In order to reveal such alterations we applied deep enzymatic fragmentation by bacterial collagenase resulting in a set of small peptides which, if modified, are likely to change their chromatographic and electrophoretic properties and can be visualized on the resulting peptide profile. Bacterial collagenase cleaves collagen mainly to tripeptides (some of them we successfuly identified).

Collagen from tail tendons of four groups of rats was studied. The tissue (a mixture of type I and III collagens) was digested by bacterial collagenase, the arising peptides were separated by HPLC and divided into five fractions. Further characterization of the profiles was done by capillary electrophoresis (CE).

It was demonstrated that the combination of HPLC and CE peptide mapping with subsequent statistical evaluation (principal component analysis and t-test) represents a reliable approach for revealing posttranslational modifications in slowly metabolized model protein in vivo.

Ekmekcioglu C¹, Haslmayer P², Philipp C¹, Mehrabi MR³, Glogar HD³, Grimm M⁴, Thalhammer T², Marktl W¹

El-Saggan AHH, Uhrík B

Polycationic dye Ruthenium red (RR) stains negative binding sites in external coats of cells, related to transport, barrier, or receptive functions, and has been used as a marker of glycoconjugates in cells of different origin. However, RR penetration into deeper parts of tissue or cells is limited due to repulsive forces of those RR⁶⁺ polycations already bound to cellular matrix. This problem may severely limit the use of RR in studies of staining properties both of tissues and of isolated cells concentrated by centrifugation. In the present study a new method of RR staining of negative binding sites on cryosections of glutaraldehyde-fixed frozen cells has been introduced with the aim to expose simultaneously all the cells and their components to the cationic dye treatment.

The mouse leukemic cell line L1210 and rat heart muscle cells from a left ventricle were used. Cells were fixed with 2% glutaraldehyde in cacodylate buffer (CB), soaked in 2.2 mol/l sucrose and frozen by plunging into liquid nitrogen. Ultrathin cryosections were cut at a temperature of -90°C, transferred with a wire-loop to Formvar coated copper grids, postfixed with 1% OsO₄ and stained with 0.05% RR in CB for 60-120 min. After removing RR solution with filter the grids were dried and examined electron microscopically.

The resulting staining was a combination of a negative contrast (the plasma membrane and membranes of intracellular organelles) and of a positive contrast (cytoplasmic matrix and the extracellular coat).

Ficková M, ¹Kotyzová D, ¹Mičková V, ¹Eybl V, Brtko J

Cadmium (Cd) and mercury (Hg) are inorganic toxicants of great environmental and occupational concern. Moreover, chronic exposure of rats to oral Cd exerts diabetogenic effect characterized by hyperglycemia and lower glucose stimulated insulin release. This study was performed to investigate the influence of toxic environmental contaminates — Cd and Hg on insulin receptors in isolated fat cells.

Male Wistar rats were drinking ad libitum either CdCl₂ (9,7 mg/l) or HgCl₂ (11,5 mg/l) in tap water for 6 weeks. This intake did not influenced weight gain (C=+237 g, Cd+255 g, Hg=+239 g). Cd treatment induced slight increase in glycemia (6,0±0,2 mmol/l) as compared to control rats (5,4±0,2 mmol/l), p<0,05. Elevated insulinemia in Cd group (0,678±0,1 ng/ml) was not statistically different either to control (0,47±0,05) or Hg (0,45±0,09) group. The number of insulin receptors (IR) in adipocytes was significantly lowered in both treated groups (C=22,3±3,6x10³/cell, Cd=9,4±1,1x10³, p<0,01; Hg = 11,1±0,9x10³, p<0.05). Despite the similar fat cell size in all groups, significantly smaller density of IR in both, Cd and Hg rats was observed. Less insulin receptors (Cd) displayed higher apparent affinity binding constant K_a (p<0,05 vs. C) and increased ED₅₀ values for insulin. A significant negative correlation (r=-0,89, p<0.001) between insulinemia and insulin receptors in isolated adipocytes of Cd group was present.

The results demonstrate the presence of IR disturbances in adipocytes as the effect of long-term intake of toxic elements.

Fi_árková B, Vytá_ek R, Vízek M,

Turanlahti et al. recently showed that hyperoxia attenuated free radical-mediated effects of NO inhalation. To test whether hyperoxia could similarly attenuate effects of endogenously produced NO, we measured nitrotyrosine concentrations in serum of rats exposed to hyperoxia during experimental pneumonia. 34 Wistar male rats were assigned to one of 4 groups. In groups 1 and 2 0,5 ml of 0,7 % carragheenan was applied intratracheally (carragheenan groups). Groups 1 and 3 were then placed for 48 hours into hyperoxia (F_IO₂ 0,75 — 0, 83) (hyperoxic groups). Rats of groups 2 and 4 breathed air (normoxic groups). 48 hours later were all rats anesthetized, intubated, their ventilation measured, their expired air collected for NO concentration measurement and 1 ml of blood taken to examine serum nitrotyrosine concentration (ELISA). ANOVA was used for statistical evaluation.

Carragheenan installation combined with hyperoxia increased NO production (159,8 ± 36,9 ppb/min compared to 22,6 ± 6,6 in controls, p< 0,05). Nitrotyrosine concentrations were in both hyperoxic groups lower than in normoxic (carragheenan-hyperoxic group 5,1 ± 0,3 uM, caragheenan-normoxic 8,3 ± 0,7; p< 0,05, control-hyperoxic 4,5 ± 0,9 and control-normoxic 7,5 ± 0,7; p< 0,05). The results suggest that hyperoxia prevents nitration of plasmatic proteins.

Turanlahti et al.: Nitric oxide and hyperoxia in oxidative lung injury. Acta Paediatr., 89, 996 -70, 2000.

Franěk M¹, Vaculín _¹, Hess L², Rokyta R¹

Medetomidin (ME), a₂ agonist, has been used in veterinary medicine for a long time to evoke sedation or as a component of combined anaesthesia. In present paper the effects of intramuscular ME alone and its combination with ketamine (KE) on pain threshold in rats are described.

ME was administered to Wistar male rats weighing 200- 230g in the doses: 100-1500 mg/kg, its effect was compared with two control groups (saline; morphine 5 mg/kg). KE (25 mg/kg) was administered alone and in combination with ME (100 and 200 mg/kg). Plantar test and tail flick were used to measure pain thresholds before, 10 and 20 min after the administrations. These two methods partially enable to distinguish spinal and supraspinal antinociceptive effects of ME.

In plantar test me evoked a shortening of response latencies for the doses 100-500 mg/kg and had no effects for the doses 750-1500 mg/kg. In tail-flick test, me evoked dose dependent antinociception for the doses over 500 mg/kg. The combination of me 100 mg/kg and ke 25 mg/kg evoked strong antinociception, 200 mg/kg me and 25 mg/kg ke evoked total anaesthesia. Ke alone (25 mg/kg) had no effect on nociception.

The obtained results indicate, that me affects nociception at two levels and in different-opposite manner: supraspinally facilitates and spinally inhibits. It is suggested that the supraspinal facilitation results from the inhibition of a tonic antinociceptive effect of bulbar nuclei (i.e. Disinhibition) while the spinal inhibition is caused by a decrease of the activity of the dorsal horn neurons participating in the nociceptive pathways.

Friedl R¹, Moeslinger T¹, Kopp B² and Spieckermann PG¹

1Institute of Physiology, Vienna, Austria

The human retina offers the unique possibility to study erythrocyte and leukocyte movement in vivo with two independent methods. The blue field entoptic technique assesses leukocyte movement in the perifoveal retinal capillaries. Using laser Doppler velocimetry and fundus photography erythrocyte movement in larger retinal vessels can be quantified. Little is, however, known about possible interactions between erythrocytes and leukocytes in the human retina. We investigated erythrocyte and leukocyte flow in the retina during administration of granulocyte colony-stimulating factor (G-CSF) under basal conditions and during states of pronounced vasoconstriction induced by systemic hyperoxia. After 8 hours G-CSF increased total leukocyte counts four-fold. This increase in circulating leukocytes was reflected by an increase in retinal white blood cell density (110 ± 48%). By contrast, neither retinal vessel diameters nor red blood cell velocity were altered by administration of G-CSF. During systemic hyperoxia a pronounced decrease in red and white blood cell flux was observed. 8 hours after G-CSF administration the reduction in white blood cell flux during systemic hyperoxia was comparable to that under baseline conditions. By contrast, the response in erythrocyte flow during systemic hyperoxia was more pronounced when G-CSF was pre-administered. These observations appear to be related to the "train" effect, a phenomenon which describes the complex interactions of leukocytes and erythrocytes in capillaries and smaller arterioles and venules. Our observation also indicates that leukocytes play an important role in retinal vascular resistance.

Gaba_ová E¹, Béder I¹, Babinská K¹, Béderová A³, Tureck_ L², Uhlíková E²

1Instute of Physiology and ²Institute of Medical Chemistry and Biochemistry, Medical School, Comenius University, ³State Health Institute, Bratislava, Slovakia

This study was designed to identify the beliefs, motivation and personal and enviromental influences shaping eating habits of a group of college students in Slovakia. We studied 167 students of medical faculty- 43 men and 124 women, average age 21.5years, who provided information on demographic and socio-economic variables, responded to an interviewer-administered, food-frequency questionnaire that assessed the consumption of more than 100 food items and 24-hours recalls of food intake. Study included measuring of blood pressure, anthropometric and lipid parameters as well as lipid peroxidation levels- conjugated diens. The prevalence of overweight was only 4% in this study, more often in women. Higher level of total cholesterol has been found in 26% of students, high levels of LDL cholesterol in 13,6%. Almost a half of students did not take food regularly, in most of the cases they replaced the main meal with the fast food. One third of the investigated group, takes vitamin and mineral supplements. However, there were variations between individuals, with specific practices being influenced by personal food preferences, time availability, health beliefs and concern, food availability, and the physical and social enviroment. Results indicate that, in general, the study group was reasonably well nourished. However, fat consumption was 30% higher than the recommended intake, for both males and females. The percentage of energy derived from carbohydrates was below the guideline value in both sexes. Relatively low iron and fiber intakes were found for females. Even though all students participated in some physical activity, less females participated in high activity sport. Based on these results, some concern about the dietary habits and the related health consequences in medical students appears justified.

The coupling between increased work and metabolism, which has been postulated for the brain more than 100 years ago, has been demonstrated now for many different tissues. In the retina, this coupling between augmented neuronal activity and increased blood flow exists as well. Using diffuse luminance flicker for photic stimulation this has been verified in several animal and human studies. A variety of techniques have been employed to show that diffuse luminance flicker induces retinal and optic nerve head vasodilatation. However, despite may efforts, the mechanism behind the coupling between increased neural activity and augmented blood as well as possible mediators involved in this process are still a matter of controversy. New knowledge on this coupling is now being accumulating thanks to the development of different techniques as laser Doppler flowmetry and new methods for exact determination of retinal vessel size, which allow the accurate assessment of the activity induced blood flow response. This presentation will provide an update on our current understanding of the coupling between increased neural activity and ocular blood flow and the factors which may potentially interfere with it. Furthermore a short overview of the techniques currently available for the assessment of neurovascular coupling will be given.

Gerová M, Smie_ko V, Kristek F

The huge body of data dealing with the role of arginine metabolism and/or NO production in genetic hypertensive rats (SHR) provide 3 possibilities: (i) NO production increase, (ii) decrease and (iii) no change. The aim of the study was to compare the response of systemic blood pressure (BP) to activators of NO synthase (acetylcholine - Ach and bradykinin - BK) in SHR and NO deficient rats (NODH). BP in age matched, anaesthetized Wistar control rats (n=8), SHR (n=7) and NODH (n=8) was 137.0 ± 2.7 mmHg, 177.1 ± 5.4 mmHg (P<0.05, vs controls), and 166.0 ± 2.7 mmHg (P<0.05, vs controls), respectively. BP decrease induced by Ach i.v. (10 µg/0.1 ml physiol. sol./10sec) amounted in controls 63.8 ± 3.2 mmHg, in SHR 108.1 ± 6.4 mmHg (P<0.05, vs controls), and in NODH 107.8±4.9 mmHg (P<0.05, vs controls). BP decrease induced by BK (100µg/0.1ml/10sec) amounted in controls 53.3 ± 5.2 mmHg, SHR 106.6 ± 8.3 mmHg (P<0.05, vs controls), and NODH 105.2 ± 4.5 mmHg (P<0.05, vs controls).

Significant increase in hypotensive response of SHR and NODH to both Ach and BK vs controls was found. No difference in rate of hypotension was found in two models of hypertension. The later finding might indicate either (i) an increased NOS activity in SHR, or (ii) a common mechanism, however different from arginine ® NO pathway, in both SHR and NODH.

Hamar J,Turchányi B, Tóth B, Vendégh Z

Aims: Ad and C fibers are responsible for antidromic vasodilatation and neurogenic inflammation (Szolcsányi J. Agents and Actions 23:4-11. 1988.). In the present studies we wanted to see whether the inactivation of these fibers could influence reperfusion injury of the skeletal muscle.

Methods: Sensory fibers of the left sciatic nerve were selectively damaged with capsaicin treatment in the anesthetized rat. 7 days later the effected hind limb underwent tourniquet ischemia for 30, 60, and 120 min. Then the limb was allowed to be reperfused for 1, 24, 72, and 168 hours. The isometric tetanic contraction force was measured of the extensor digitorum longus muscle (EDL) on both sides (damaged vs. control). Contractions were elicited with direct and indirect electric stimulations of the EDL and the sciatic nerve, respectively.

Main results: Capsaicin alone does not effect the EDL contraction force. Selective denervation did not influence the contractions after 30 min ischemia. After 60 min ischemia the selectively denervated muscle contracted slightly weaker at 168 hours of reperfusion. Contraction force of the capsaicin treated side was significantly stronger at all reperfusion times and stimulation modes.

Conclusions: A 2-hour ischemia produces qualitatively more serious damages. Exclusion of the peptidergic sensory afferents can reduce the inflammatory reaction, a major component of reperfusion injury.

Herget J¹ , Novotná J, Bláhová L¹

Hypercapnia attenuates the hypoxic pulmonary hypertension (HPH) and oxygen radical injury induced by exposure to chronic hypoxia . HPH results from remodeling of peripheral pulmonary arteries. We hypothesize that the remodeling is induced by matrix collagen cleavage in the vascular wall . Collagen composition of peripheral pulmonary arteries was studied in male adult rats exposed to 4 days chronic isobaric hypoxia (F_iO2 = 0.1, F_iCO2 = 0, n = 7), hypoxia and hypercapnia (F_iO2 = 0.1, F_iCO2 = 0.045, n = 7) and in normoxic controls (n = 7). Collagenous proteins were extracted from the isolated peripheral pulmonary arteries and analyzed by SDS electrophoresis . Similarly to our previous study in all hypoxic rats, we detected the 3/4 and 1/4 collagen type I cleavages. Cleavages were not present in rats exposed to normoxia and in rats exposed to hypoxia combined with hypercapnia.

We conclude that hypercapnia attenuates HPH by inhibition of collagen cleavage in the walls of peripheral pulmonary arteries. The mechanism is probably related to the CO₂ inhibition of hypoxia induced radical tissue injury.

Hijová E¹, Ni_tiar F², Petrá_ová D¹

Mercury is a serious ubiquitous pollutant (environmental and occupational), which have been reported as potent toxic and/or carcinogenic agens in humans and animals. Mercury enters an organism in a variety of chemical forms, causes the production of reactive ogygen species leading to the formation of an oxidative stress.

Methods: Adult male Wistar albino rats (n=60, 10 per group) were for 30d. fed on a normal laboratory diet and supplied with drinking water containing mercury (as mercuric chloride) in different doses of LD₅₀. The LD₅₀ for HgCl₂ is 37 mg.kg^-1. The rats in control group received drinking water without mercuric chloride. Plasma specimens were used for determination of vitamin C as antioxidant and malondialdehyde as product of lipid peroxidation and stress indicator.

Results: In the group receiving dose LD₅₀ of mercuric chloride the concentration of ascorbic acid and malondialdehyde were increased by 22.19% and by 6.88% respectively, compared with control group. Statistically significant was comparison dose of LD₅₀ of HgCl₂ with 1/4 of LD₅₀ for ascorbic acid (p<0.01), and for malondialdehyde (p<0.001).

Hinghofer-Szalkay H, ¹Jezova D, Loder I, Roessler A

Purpose of our study was to identify stimulus combinations of lower body negative pressure (LBNP) and head-down tilt (HDT) which compensate each other in their effect on cardiovascular variables, electrical thoracic impedance, hematocrit and plasma mass density, and hormone concentrations in blood plasma. We hypothetized that „neutral points" (NP’s) can be demonstrated (hypothesis 1); NP’s may vary with variables (hypothesis 2); and NP’s might change as a function of time (hypothesis 3). Tests were performed in 10 healthy young subjects. Normalized values were used to compare with supine control conditions SC to get rid of time-dependent shifts unrelated to the stimulus. The effect of a certain stimulus pair X at time t was defined as the normalized value at(X,t) minus the normalized value at (SC,t). Data were plotted as a function of tilt angle, data points connected by a best-fit line or curve, and the NP determined as inter- or extrapolated NP’s, defined as the intersection with the respective supine control line. Preliminary findings suggest that with LBNP-15, the NP is 13-16° HDT for thoracic impedance (irrespective of stimulus duration), and plasma renin activity (PRA), and >24° for aldosterone; no clear NP was found for heart rate, blood pressure, plasma density, and hematocrit. With LBNP-35, the NP is about 16-30° for heart rate (increasing with stimulus duration), 26-28° HDT for PRA and aldosterone, and Ò30° HDT for thoracic impedance, plasma density, and hematocrit. It might be concluded that NP’s do not always clearly show (refutes H1), that NP’s differ for different variables (confirms H2), and there is a dependency on stimulus duration in some instances (partially confirms H3).

Hitka P, Vízek M

Analyses of expired air became an important resource of clinical information (1). Such analyses require relatively large volumes of expired air therefore, in studies using small experimental animals, long lasting collection of expired air is necessary. This paper describes a system exhausting expired air of spontaneously breathing anesthetized, intubated rats.

Tidal volume of anesthetized, intubated rat was recorded by classical body plethysmography. The volume signal was used to trigger pump, sucking air from the side port of tracheal cannula. Onset of either inspiration or expiration activated an electronic controller regulating the time delay with which the pump was switched on and intensity of the sucking. The flow sucked by the pump resembled the expiratory flow of the animal.

The system was tested by comparing CO₂ output measured by collecting the expired air by our system with classical collection of expired air. The differences between the methods ranged from +0,2 to —0,1 ml/min. The respective CO₂ outputs (1,6 ± 0,4 and 1,5 ± 0,4 ml/min (mean ± SD)) did not significantly differ.

References:

1. Mutlu, G.M. et al.: Collection and analysis of exhaled breath condensate in humans. Am J Respir Crit Care Med, 2001, 164, 731-737

Hlavacka F, Polonyova A

Postural instability in cosmonauts returning from spaceflight probably results from in-flight adaptation of central nervous system processing of sensory inputs from vestibular, somatosensory and visual systems. We tested how post-spaceflight postural reactions to unilateral vibration of lower leg muscles (somatosensory input) were changed. For comparison the postural responses to soleus and tibialis anterior muscles vibration with frequencies (40-100 Hz) were measured in healthy subjects.

Vibration (duration 8s, amplitude 1 mm and frequency 80 Hz) was applied to the belly of tibialis anterior (TA) or soleus (SOL) muscle. The force platform measurements (center of foot pressure - CoP) were performed before spaceflight and on the first day (10 hours) after landing, the second day and the fifth day.

Unilateral vibration of the TA muscle induces tilt of body forwards and in the direction of the stimulated leg. While unilateral vibration of SOL muscle causes a backward tilt and in the direction opposite to the stimulated leg. In the first days after spaceflight an increased postural responses to soleus vibration were observed. Influence of microgravity on postural response to TA vibration was negligible. Comparison of body responses to the left and to the right leg vibration showed changes in proprio-postural asymmetry. The balance recovery was successfully finished after 5 days.

The results showed that the magnitude of postural responses (body tilt) to leg muscle vibration was modulated by the frequency of the vibratory stimulus. The frequency of vibration does not influence direction of body tilt, which was determined only by the muscle stimulated.

Impairment of the ocular perfusion represents a major pathophysiological mechanism for the development of several ocular diseases including open-angle glaucoma, which is one of the leading causes of blindness. Endothelin-1 has an important role in the regulation of the optic nerve head and retinal blood supply, systemic blood pressure as well as aqueous humour outflow and consequently regulates the intraocular pressure. These factors in combination determine the ocular perfusion pressure and its diurnal fluctuation, which seem to be key mechanisms in the development and progression of the retinal ganglion cell apoptosis in open-angle glaucoma. Animal models for the endothelin induced damage of the optic nerve head perfusion, decrease of the aqueous humour outflow and elevation of the intraocular pressure as well as low systemic blood pressure as a consequence of low plasma endothelin-1 concentration in the human will be shown in the lecture. The clinical consequences for glaucoma will be also discussed.

Honzíková N¹, Hrstková H², Hak J³, Balcárková P¹, Závodná E¹, Nováková Z¹ 

Balance between the reflex and tonic autonomic control of the heart in children with bronchial asthma was studied.

Blood pressure was recorded in 15 children (mean age±SD: 12.9±1.5 years, range 11-15 years) with diagnosis of bronchial asthma at the beginning (group A1) and after 6 weeks of balneal therapy (group A2) and in 45 healthy controls (C), age and sex matched (spectral