Janette Dias Souza

GABA and specific GABA binding sites in brain nuclei associated with vagal outflow

Abstract GABA levels and specific (3H) GABA binding were determined in several nuclei of cat brain. Since previous pharmacological studies (DiMicco et al. [3]) suggested that nucleus ambiguus (NA) may be the site of a GABA-receptor mediated inhibition of vagal outflow to the heart, we were interested in comparing the GABA content and density of (3H) GABA binding in NA with that of other nuclei known to contain GABAergic synapses. The GABA content of NA was 21.2 ± 2.4 nmol/mg protein, similar to that found in the caudate nucleus (28.4 ± 2.9 nmol/mg protein) and 2.5 fold higher than the GABA content of the surrounding reticular nuclei. In frozen-thawed and Triton X-100 treated membranes prepared from NA, specific GABA binding was 98 ± 28 fmol/mg protein when measured using 30 nM (3H) GABA. This was more than 3 fold higher than binding obtained in surrounding reticular tissue and approximately half the value obtained in substantia nigra. GABA content of paired right and left NA was nearly equal; however specific GABA binding of paired right and left NA differed markedly, with the right NA usually exhibiting greater specific binding than the left NA. Retrograde degeneration of vagal fibers of NA by intracranial sectioning of the right vagal trunk decreased the asymmetry in GABA binding of paired right and left NA. Asymmetry was also noted in the percent of the reflex-induced bradycardic response mediated by each vagus nerve. These results suggest that GABAergic synapses may be present in NA, and that some of the postsynaptic receptors for GABA may be associated with vagal efferents. The bilateral asymmetry in the physiological reflex-induced response coupled with the bilateral asymmetry in GABA binding in NA suggests that the degree of vagal activity emanating from NA may be determined by the density of GABA receptors.

Hypotensive effect of urapidil: CNS site and relative contribution.

Summary: The purposes of our study were to determine the contribution of the CNS to the hypotensive effect of urapidil in the cat and the specific brain site of action of this agent. For the first purpose, urapidil was studied on preganglionic sympathetic nerve activity, arterial pressure, and heart rate. Three systemic bolus doses of urapidil were administered (0.22, 0.44, and 1.3 mg/kg). All three doses lowered arterial pressure, and the highest dose produced a significant decrease in sympathetic nerve discharge in five of six animals studied. The lower two doses had no significant effect on sympathetic activity, and none of the doses altered heart rate. These results suggest that a high i.v. dose of urapidil is required to evoke hypotension by an action in the central nervous system (CNS). For the second purpose, urapidil was applied bilaterally to the intermediate area of the ventral surface of the medulla in doses of 25 and 50 µg. These doses caused decreases in arterial pressure of −6.1 ± 2.2 (p < 0.05) and −21.0 ± 5.9 (p<0.05) mm Hg, respectively, but no change in heart rate. In addition, respiratory stimulation also occurred with the higher dose as respiratory minute volume increased by 81 ± 14 ml/min (p < 0.05). The highest dose of urapidil had no effect on arterial pressure when applied to other chemosensitive areas of the ventral surface of the brain. Comparative studies with prazosin (10 µg applied bilaterally to the intermediate area) indicated no hypotensive effect of this α1-adrenoceptor blocking agent. These results suggest that the central hypotensive effect of urapidil occurs at the intermediate area and does not involve blockade of α1-adrenoceptors.

Comparative Effects of Urapidil, Prazosin, and Clonidine on Ligand Binding to Central Nervous System Receptors, Arterial Pressure, and Heart Rate in Experimental Animals

We studied the effects of urapidil, clonidine, and prazosin on ligand binding to central nervous system receptors in rats and on arterial pressure and heart rate in chloralose-anesthetized cats. Ligand binding studies indicated that urapidil had 90 times greater affinity for alpha 1 than for alpha 2 adrenergic receptors. Administration of urapidil (129 micrograms) into the cerebroventricles of cats revealed no effect after lateral ventricle injection, a decrease of 9.7 +/- 3.0 mm Hg after fourth ventricle injection, and an increase of 10.8 +/- 2.2 mm Hg after restriction of the drug in the forebrain ventricles. Clonidine (30 micrograms) produced hypotension and bradycardia after injection into the lateral ventricle. Prazosin was ineffective after cerebroventricular injection. Intravenous administration of 0.22, 0.67, and 2.00 mg/kg urapidil produced dose-dependent decreases in arterial pressure that were associated with blockade of alpha 1 adrenergic receptors. Intravenous administration of prazosin elicited the same response. Clonidine (10 micrograms/kg, intravenously produced an initial increase in arterial pressure that was unaffected by pretreatment with urapidil or prazosin. These results suggest that urapidil produces hypotension by an action on the peripheral vasculature and in the hindbrain. The peripheral effect involves blockade of alpha 1 adrenoceptors.

Chemical stimulation of the area postrema induces cardiorespiratory changes in the cat

The purpose of our study was to determine the cardiorespiratory effects of exciting cell bodies of the area postrema of the cat. This was accomplished by local application of L-glutamic acid (bilateral application of 5 microliter of a 250-1000 mM solution) and kainic acid (bilateral application of 5 microliter of a 40 mM solution) to the area postrema of chloralose-anesthetized cats while monitoring arterial pressure, heart rate, tidal volume and respiratory rate. These excitatory amino acids activate neuronal cell bodies but not axons of passage. L-Glutamic acid produced a dose-dependent increase in arterial pressure, decreases in respiratory rate and minute volume and, occasionally, ventricular tachyarrhythmias. Kainic acid produced effects similar to those seen with L-glutamic acid except the changes in respiratory activity were more pronounced with each animal exhibiting respiratory arrest. In artificially respired animals, kainic acid produced similar cardiovascular changes as those occurring in spontaneously breathing animals (i.e. increases in arterial pressure of 61 +/- 5.7 mm Hg, and in heart rate of 32 +/- 8.3 beats/min). Finally, application of kainic acid to the area postrema abolished the pressor and tachycardic responses to bilateral occlusion of the carotid arteries. These results suggest that activation of cell bodies in the area postrema can result in pronounced cardiorespiratory changes.

Evidence for a role of central serotonergic neurones in digitalis-induced cardiac arrhythmias.

EVIDENCE indicates that digitalis drugs administered intravenously increase central sympathetic outflow and that this in turn results in cardiac arrhythmias1–6. The central nervous system transmitter(s) that mediates this effect is not known. According to Saito et al., noradrenaline, the primary transmitter studied so far, is not involved in the case of guinea pigs7. 5-Hydroxytryptamine (5-HT) however, may be an important chemical mediator regulating central sympathetic outflow8,9, and it is involved in the respiratory arrest induced in rats by intravenous administration of digitoxigenin10. We now report evidence that suggests that brain 5-HT is involved in digitalis-induced cardiac arrhythmias.

Increase in coronary vascular resistance produced by stimulating neurons in the region of the area postrema of the cat

Recently, Somberg, in a preliminary report (1983), noted that electrical stimulation of the area postrema causes an increase in coronary vascular resistance. The increase in resistance was mediate by an increase in sympathetic activity as it was counteracted by alpha-adrenergic receptor blockade. The purpose of our study was to confirm the findings of Somberg by using the method of chemical-induced activation of cell bodies in the area postrema. This was done by bilateral topical application of kainic acid to the area postrema of cats while monitoring coronary blood flow, heart rate, arterial blood pressure and the ECG. Topical application of kainic acid produced an increase in coronary vascular resistance (43 +/- 13%, P less than 0.05), S-T segment changes, increases in heart rate (60 +/- 10, P less than 0.05) and arterial blood pressure (88 +/- 20, P less than 0.05) and ventricular tachyarrhythmias. All of these effects were prevented by pretreating animals with the alpha-adrenoceptor blocking agent, phentolamine. These results suggest that chemical excitation of area postrema neurons produces coronary constriction that is mediated by the sympathetic nervous system and alpha-adrenoceptors on coronary vessels.

Stress-induced changes in the function of the parasympathetic nervous system are mimicked by blocking GABA in the CNS of the cat

The purpose of this study was to determine the effect of blockade of receptors for gamma-aminobutyric acid (GABA) in the forebrain, on vagal activity to the stomach and heart. This was done by injecting bicuculline (50 micrograms) into the lateral ventricle of the brain and restricting the drug to the forebrain ventricles by cannulating the cerebral aqueduct. Studies were performed in chloralose-anesthetized cats and gastric motility was monitored using extraluminal force transducers, sutured to the antrum and pylorus. Cardiac vagal activity was determined by noting the sinus bradycardia that developed from activation of the baroreceptor reflex induced by phenylephrine. Administration of bicuculline into the lateral ventricle of 7 animals produced increases in the minute motility index of 5.3 +/- 0.8 (antrum) and 13.9 +/- 2.1 (pylorus). This was associated with inhibition of baroreceptor-induced vagal bradycardia (i.e. -38 +/- 6.4 beats/min before bicuculline and -7.7 +/- 5.7 beats/min after bicuculline). These data indicate that a GABAergic mechanism in the forebrain may be important for controlling vagal outflow to both the stomach and the heart.

Deleterious effects of taurine in cats with digitalis-induced arrhythmias

An established model of digitalis toxicity was used to investigate the antiarrhythmic properties of taurine. I.v. doses of taurine ranging from 0.01 to 4.0 mmole/kg were ineffective in converting a deslanoside-induced arrhythmia to sinus rhythm. Indeed, taurine was found to aggravate the arrhythmia and in three experiments precipitated ventricular fibrillation. In addition, pretreatment with 5 mmole/kg taurine i.v. had no significant effect on the doses of deslanoside to produce ventricular arrhythmia and fibrillation.

Interaction of serotonin and deslanoside on cardiac rhythm in the cat.

The present study was performed to determine whether increases in the tissue content of serotonin creatinine SO4 in the periphery would influence the arrhythmogenic effect of deslanoside. This was accomplished by infusing serotonin into anesthetized cats exposed to a subarrhythmic dose of deslanoside, determining doses of deslanoside required to produce ventricular tachycardia and ventricular fibrillation, and determining ventricular pacemaker rate (obtained during vagal-induced sinus node suppression). It was found that animals receiving serotonin creatinine SO4 plus deslanoside exhibited a greater increase in ventricular rate during sinus node suppression than with 5-HT infusion alone. No corresponding increase in ventricular pacemaker rate during sinus node suppression was observed with creatinine SO4 plus deslanoside. In addition, the dose of deslanoside to produce ventricular fibrillation in these animals was significantly correlated with the increase in ventricular pacemaker rate seen during th 5-HT infusion in the presence of deslanoside. Studies were also performed to determine whether the arrhythmogenic interaction of serotonin with deslanoside was associated with alterations in either cardiac tissue, blood or plasma levels of serotonin and 5-hydroxyindoleacetic acid. The data revealed a significant correlation between serotonin content in the left ventricle and the dose of deslanoside required to produce ventricular fibrillation. These results suggest that exogenous serotonin interacts with deslanoside to enhance the arrhythmogenic action of deslanoside.

Blockade of bicuculline-induced pressor and tachycardic responses by forebrain administration of muscimol

To determine whether i.v. administered bicuculline acts in the forebrain to increase arterial blood pressure and heart rate, this agent was administered i.v. to chloralose anesthetized cats that had muscimol injected into and restricted to the forebrain ventricles. Bicuculline (0.5 mg/kg i.v.) given alone increased arterial pressure by 56 +/- 8 mm Hg and heart rate by 45 +/- 11 beats/min. Bicuculline given to animals exposed to muscimol exhibited no increase in either of these parameters. Muscimol localized to the forebrain did not alter the pressor response to a non-GABA antagonist agent, strychnine, indicating a specific interaction of the drugs with GABA receptors in the forebrain.