Stanislav Trojan

Correlation between electrocorticographic and motor phenomena induced by pentamethylenetetrazol during ontogenesis in rats

Motor and electrocorticographic (ECoG) phenomena induced by pentamethylenetetrazol (PTZ) were directly compared in adult as well as in young rats (age groups of 7, 12, and 18 days). The technique for implantation of cortical electrodes was elaborated in young rats. Myoclonic jerks were elicited by PTZ in all age groups studied. In adult animals they were accompanied by isolated spikes and/or spike-and-wave complexes, whereas in 7-day-old rats there was a poor correlation between ECoG abnormalities and myoclonic jerks. With increasing age an improved correlation between ECoG abnormalities and behavioral changes were noted. After further injections of PTZ, rats had generalized tonic-clonic motor seizures. Ictal ECoG activity usually started in one cortical region in 7-day-old rats and became progressively generalized whereas in older rats generalized electrical discharges accompanied the motor seizures from onset. The clonic phase of motor seizures was not always accompanied by specific ECoG changes in young rats. At all ages the correlation between ECoG activity and individual phases of the generalized tonic-clonic seizures was poor.

Ontogenetic development of isonicotinehydrazide-induced seizures in rats.

Motor seizures elicited by isonicotinehydrazide (INH) were studied in 188 male albino rats aged from 5 days to adulthood. INH was found to be able to induce both minimal, predominantly clonic, and major, i.e. generalized, tonic-clonic seizures during the whole course of development. Tonic hindlimb extension was observed as a part of major seizures even in the youngest group studied. Fifty-percent convulsant doses counted in 7-, 12-, 18-, 25- and 90-day-old rats did not significantly differ. The developmental changes found were a poor time resolution of minimal and major seizures during the first two postnatal weeks and significantly longer latency of major seizures in 5- and 7-day-old rat pups in comparison with all other age groups. The importance of the developmental stage of a generator of seizures as well as of the maturation of triggering mechanisms was demonstrated comparing the present results with our previous data.

Blphasic changes in the density of muscarinic and beta-adrenergic receptors in cardiac atria of rats treated with diisopropylfluorophosphate

Chronic treatment with organophosphate inhibitors of cholinesterases is known to bring about down-regulation of muscarinic acetylcholine receptors in the heart while its effect on the functionally antagonistic beta-adrenergic receptors is not known. We describe experiments in which rats were exposed to daily injections of diisopropylflurophosphate (DFP) and the density of muscarinic and beta-adrenergic binding sites in their cardiac atria was measured according to the binding of (3H)quinuclidinyl benzilate ((3H)QNB) and (-)-4-(3-tert-butylamino-2-hydroxy)-propoxy-(5,7-3H) benzimidazol-2-one ((3H)CGP 12177) as subtype non-specific muscarinic and beta-adrenergic ligands, respectively. Biphasic course of changes was discovered with both ligands. With the dosage scheme applied, the density of beta-adrenoceptors was augmented 24 h after the first dose of DFP and decreased to about one half of control values after 5 days of treatment with DFP. The density of muscarinic receptors was augmented after two days of treatment with DFP and decreased to about one half of control values after 5 days of treatment. Comparatively small changes in the heart rate were observed during the treatment, they reflected changes in the muscarinic and beta-adrenergic receptor density. The finding of DFP-induced changes in the binding of (3H)CGP 12177 suggests that the mechanisms responsible for the control of the density of muscarinic and beta-adrenergic receptors in the heart cells are interconnected but potential roles of other factors involved in in vivo experiments deserve further analysis. The transient increase in the density of muscarinic receptors after two days of DFP treatment appears related to published data on transient stimulation by cholinergic agonists of the transcription of mRNAs for muscarinic receptors.

EFFECTS OF SINUSOIDAL MAGNETIC FIELD ON ADHERENCE INHIBITION OF LEUKOCYTES

Response of leukocytes to exposure to an external magnetic field with frequency 50 Hz and sinusoidal waveform was investigated in vitro using the leukocyte adherence inhibition (LAI) assay developed as a measure of cell-mediated immunity. Leukocytes taken from healthy humans adhere, but their adherence decreases after 1 hr of exposure to the magnetic field with magnetic induction of 1 and 10 mT. The majority of leukocytes taken from cancer patients before any medical treatment do not adhere, and exposure to the magnetic field increases adherence. Correlation between the LAI assay results and the cell-mediated immunity suggests an effect of magnetic fields on leukocyte immune function in humans.

Heterologous regulation of muscarinic and beta-adrenergic receptors in rat cardiomyocytes in culture

Previous work indicated that hyperstimulation of muscarinic receptors brings about profound changes not only in the density of the muscarinic receptors, but also of the beta-adrenoceptors in rat heart atria in vivo. We have now investigated whether a similar receptor cross-regulation occurs in cardiomyocytes in vitro. Cardiomyocytes from 3-4 day old rats were exposed to chemical agents on days 5-6 in culture. Densities of muscarinic and beta-adrenergic receptors were measured according to the binding of N-[3H]methylscopolamine and [ H]CGP 12177, respectively, to cell surface membranes and cell homogenates. Exposure of cells to the muscarinic agonist carbachol (1 mmol/l) brought about a profound decrease in the number of muscarinic receptors. The number of beta-adrenoceptors displayed biphasic changes, being augmented after 24 h (by 20-45% on the cell surface and by 29% in the homogenate) and diminished after 48 h and 72 h (after 48 h, decrease by 44-75% on the cell surface and by 36% in the homogenate). These effects of carbachol were not prevented by dimethylaminopropyl-bis-indolylmaleimide, the inhibitor of protein kinase C. Exposure of cells to the beta-adrenoceptor agonist isoprenaline (0.1 mmol/l) strongly diminished the number of beta-adrenoceptors on the cell surface and in the homogenate. The density of muscarinic receptors on the cell surface was diminished by 24-43% after 24 h exposure to isoprenaline and unchanged after 48 h, whereas the concentration of muscarinic receptors in the homogenate was unchanged after 24 h and increased by 20% after 48 h. The isoprenaline-induced decrease in the density of cell surface muscarinic receptors could not be simulated by forskolin and was not abolished by the protein kinase A inhibitors Rp-cAMPS and HA-1004. Dibutyryl cyclic AMP diminished the density of cell surface muscarinic receptors more than that of the beta-adrenergic receptors. Our data reveal a novel phenomenon of a biphasic change (an increase followed by a loss) in the density of beta-adrenoceptors during exposure of cardiocytes to carbachol. Activation of beta-adrenoceptors brings about less conspicuous changes in the density of muscarinic receptors. The observed phenomena of receptor cross-regulation cannot be explained by simple activations of protein kinases A and C.

Effects of Magnetic Field 0.1 and 0.05 mT on Leukocyte Adherence Inhibition

T lymphocytes taken from healthy humans and cancer patients before and after medical treatment were exposed to the magnetic field 0.1 and 0.05 mT to study response of the cell-mediated immunity. Leukocyte adherence, which is considered to correlate with the cell-mediated immunity, was measured using an in vitro technique—leukocyte adherence inhibition (LAI) assay. Exposure to the magnetic field increases adherence of T lymphocytes especially those from cancer patients before medical treatment. The effects of exposure to the magnetic field 0.1 and 0.05 mT are similar to those of greater magnetic fields in the range of 0.5 to 10 mT. The effects of the AC and DC magnetic fields 0.05 mT do not display large differences attributable to the magnetic field.

Effects of sinusoidal magnetic field on adherence inhibition of leucocytes: preliminary results

The leucocyte surface properties manifest the cell-mediated immunity. The response of the cell-mediated immunity to external magnetic field was examined by observing leucocyte adherence to solid state surfaces. In the presence of antigen, leucocytes taken from cancer patients exhibit decreased adherence in contrast with adherence of leucocytes from healthy humans. After 1 h exposure to a sinusoidal magnetic field of 50 Hz and of 1 mT or 10 mT, adherence of leucocytes taken from cancer patients is strongly increased. The 1 mT magnetic field has stronger effect than the 10 mT field.

Effects of short-term and prolonged aerogenic hypoxia on gamma-glutamyl transpeptidase activity in the brain, liver, and biological fluids of young rats

Posthypoxic fluctuations in the levels of two excitatory amino acids, glutamate and aspartate, may be related to changes in mechanisms(s) which are responsible for their reuptake. As gamma-glutamyl transpeptidase (GGT) plays a role in mediating the uptake of glutamate and aspartate into various compartments of the brain, we studied changes in the activity of this enzyme in main regions of the brain in young and adult rats. We found a posthypoxic increase in bound GGT activity in some brain regions of 18-day-old animals after acute exposure, but no changes were observed after prolonged altitude hypoxia, with the exception of a decrease in cortical GGT activity. In contrast, acute hypoxia decreased GGT activity in the cortical capillaries to 59%, but prolonged hypoxic exposure was ineffective. However, the activity of soluble GGT in the cerebrospinal fluid of both groups of rats was several-times elevated in comparison with controls. At the same time, bound GGT activity was increased in the liver after acute or prolonged altitude hypoxia. The soluble GGT activity in plasma was only increased after prolonged exposure. Ninety days after prolonged hypoxic exposure the bound GGT activity was reduced in all brain regions to about 60–70% of controls (significantly higher in females than in males) as long-term developmental sequel from early postnatal hypoxia.

Effects of Sinusoidal 0.5 mT Magnetic Field on Leukocyte Adherence Inhibition

Exposure of T lymphocytes to an external 50 Hz and 0.5 mT magnetic field was investigated in vitro using leukocyte adherence inhibition (LAI) assay which is a measure of cell-mediated immunity. Adherence of T lymphocytes taken from healthy humans and from cancer patients before and after medical treatment is enhanced after 1 h exposure to the magnetic field. The experimental findings for the magnetic field 0.5 mT are compared with published data for 1 and 10 mT. The results are consistent with suggestions of magnetic field effects on immune function in humans.

STUDY OF THE INTERACTION FORCES IN HUMAN CD4LY

Abstract The cluster formation of CD4Ly cells (a subpopulation of T lymphocytes) in suspension was examined. CD4Ly cells were prepared from blood taken from cancer patients and from healthy counterparts (blood donors). CD4Ly cells taken from cancer patients formed tightly bound clusters in suspension with corresponding tumour antigen or LDH (lactic dehydrogenase) virus antigen. Cluster formation was a rare event in suspensions with CD4Ly cells taken from healthy humans. Cluster formation coincides with decreased adherence of CD4Ly to solid state surfaces and suggests strong mutual interaction between CD4Ly. A tentative theoretical explanation based on the Frohlich coherent vibration hypothesis is given.