Zafer Gören

The role of amygdala and hypothalamus in GABAA antagonist bicuculline-induced cardiovascular responses in conscious rats

gamma-Aminobutyric acid (GABA) is known to play an important role in the central control of cardiovascular functions. GABAergic agonists and antagonists elicit blood pressure and heart rate changes when injected into the brain. It was demonstrated here that bicuculline methiodide (BMI), a GABAA antagonist, caused dose-dependent increases in both blood pressure and heart rate in conscious rats when injected intracerebroventricularly. The roles of the central nucleus of the amygdala (CeA), the paraventricular nucleus (PVN) and the dorsomedial nucleus (DMH) of the hypothalamus in BMI-induced blood pressure and heart rate changes were investigated in this study. The pressor effect of BMI was significantly attenuated by the electrolytic ablation of DMH and PVN, whereas it was only slightly, but insignificantly reduced by CeA lesions. The microinjection of BMI into the DMH and the PVN elicited significant pressor and tachycardic responses whereas only a slight increase was observed in rats injected BMI into the CeA. The BMI-induced increases in both blood pressure and heart rate were more prominent when given into the DMH. These results indicate that the DMH plays an important role in GABAergic control of cardiovascular functions. The PVN and CeA seem to have a minor part in this respect.

Reversal of hemorrhagic shock in rats by oxotremorine: the role of muscarinic and nicotinic receptors, and AV3V region.

In an experimental model of hemorrhagic shock resulting in the death of almost all rats within 20-30 min, centrally active cholinomimetic drugs are reported to induce a prompt, sustained and dose-dependent improvement in blood pressure and survival rate claimed to be due to nicotinic, but not muscarinic actions. In the present study, cholinergic receptor agonist, oxotremorine (50 micrograms/kg, i.v.) increased mean arterial pressure (from 22 +/- 1 to 123 +/- 3 mm Hg) and 60 min-survival rate (from 0% to 92%) in rats bled to hypovolemic shock. Atropine (2 mg/kg, i.v.) pretreatment inhibited the pressor effect of oxotremorine significantly, but did not modify its effect on survival rate. On the other hand, pretreatment with mecamylamine (50 micrograms, i.c.v.) almost abolished the reduction in mortality rate, but inhibited the pressor effect of oxotremorine, partially. These results indicate that oxotremorine-induced pressor response and decrease in mortality in rats with severe hemorrhagic shock are primarily mediated via central muscarinic and nicotinic receptors, respectively. AV3V region was previously reported to be involved in pressor and natriuretic effects of i.c.v. carbachol in normotensive rats. In the present study, the electrolytic lesions of AV3V region significantly inhibited oxotremorine-induced increases in both blood pressure and survival rate in rats subjected to hemorrhagic shock. These findings indicate that AV3V region plays a major role in cholinergic cardiovascular control in hypotensive animals as well as normotensives.

Modulation of the pressor response elicited by carbachol and electrical stimulation of the amygdala by muscarinic antagonists in conscious rats

The nature of the muscarinic receptor involved in mediating cardiovascular changes caused by unilateral microinjection of carbachol (5 nmol) into, and electrical stimulation (200–300 μA) of, the amygdaloid complex was investigated in conscious, unrestrained female Sprague‐Dawley rats. Unilateral microinjection of carbachol (5 nmol; n=6) and electrical stimulation (200–300 μA, 80 Hz, 30 s; n=4) caused a significant rise in blood pressure of 21±4 mmHg and 25±5 mmHg, respectively. These changes were associated with no overall effect on heart rate. The effects of electrical stimulation were found to be repeatable. Pretreatment i.c.v. with pirenzepine (5–20 nmol; n=6–7 for each dose), dose‐dependently inhibited the rise in blood pressure induced by carbachol, whereas AF‐DX 116 (100 nmol; n=6) failed to have any effect on the carbachol‐induced pressure response. Neither antagonist alone had any effect on resting baseline variables. Unilateral microinjections of atropine sulphate (1–100 nmol; n=4–6 for each dose), pirenzepine (0.03–10 nmol; n=4 for each dose) or AF‐DX 116 (10–60 nmol; n=4–5 for each dose), into the amygdala, dose‐dependently inhibited the rise in blood pressure caused by electrical stimulation (200–300 μA). The ID50 values were 1.05, 0.23 and 39.5 nmol, respectively. Although pirenzepine seemed to be more potent than atropine, this difference was not significant. It is concluded that the rise in blood pressure elicited by unilateral microinjection of carbachol into, or electrical stimulation of, the amygdaloid complex is mediated by M1‐muscarinic receptors.

Subtypes of muscarinic receptors in rat duodenum: a comparison with rabbit vas deferens, rat atria, guinea-pig ileum and gallbladder by using imperialine.

The specific binding of [3H]QNB to rat duodenum smooth muscle membranes was a saturable process and Scatchard transformation of the saturation curves indicated a linear plot (nH = 1.017+/-0.071). The K(D) and Bmax values were 0.168+/-0.025 nM and 46.7+/-8.6 fmol/mg protein, respectively. Analyses of competition curves using pirenzepine and guanylpirenzepine indicated more than one class of binding site. A minor population of muscarinic binding sites showed high affinity (M1) for both pirenzepine (19.3+/-1.2%; pKi = 8.29+/-0.36) and guanylpirenzepine (29.4+/-2.0%; pKi = 7.28+/-0.11). The antagonistic affinity values of pirenzepine and guanylpirenzepine for the remaining low affinity binding sites, and that of methoctramine indicated the presence of both M2 and M3 subtypes. McN-A-343 produced relaxations in rat duodenum and inhibited twitch contractions of rabbit vas deferens induced by electrical stimulation in a concentration dependent manner. Carbachol (Cch) exerted concentration-dependent negative inotropic effect in rat atria and contractile effects in guinea-pig gallbladder and ileum longitudinal muscle-myenteric plexus preparation. Imperaline displaced the concentration-response curves to McN-A-343 and Cch to the right in parallel, without affecting the maximum responses in all tissues studied. The rank order of the pA2 values was rabbit vas deferens > rat atria > guinea-pig gallbladder = guinea-pig ileum > rat duodenum. The presynaptic muscarinic receptors at the rat duodenum and rabbit vas deferens were concluded to be of M1 and M4 subtypes, respectively.

Role of paraventricular and dorsomedial nuclei of the hypothalamus and central nucleus of the amygdala on muscimol‐induced cardiovascular responses

Summary— Gamma‐aminobutyric acid (GABA) plays an important role in the central control of cardiovascular functions. Previous evidence indicates that a tonically active GABAergic system exists in forebrain structures. The purpose of this study was to examine the role of the unilateral lesion of the central nucleus of amygdala, paraventricular or dorsomedial nuclei of the hypothalamus on muscimol‐induced cardiovascular responses. Electrolytic ablation of nuclei was made by a monopolar isolated electrode under a stereotaxic instrument, 3–5 days before the experiments. Effects of intracerebroventricular injections of muscimol were investigated in intact, lesioned and sham‐lesioned rats. On the day of the experiments, blood pressure and heart rate recordings were carried out in male Sprague‐Dawley conscious rats. Muscimol produced decreases in arterial blood pressure and heart rate. The hypotensive effect of muscimol was completely inhibited in rats with dorsomedial nucleus lesions, whereas the bradycardic effect was partially prevented. The results indicate that the dorsomedial nucleus of the hypothalamus plays an important role on muscimol‐induced blood pressure and heart rate responses.

Carbachol-induced pressor responses and muscarinic M1 receptors in the central nucleus of amygdala in conscious rats

The type of muscarinic receptor in the central nucleus of the amygdala that mediates the carbachol-evoked pressor responses was investigated in conscious unrestraint Sprague-Dawley rats. Carbachol (100 ng) injected into the lateral cerebral ventricle caused a significant rise in blood pressure of 31.8+/-4.5 mmHg and a decrease in heart rate of 80.0+/-12.2 beats/min. Pirenzepine (10-75 nmol) injected into the central nucleus of the amygdala inhibited carbachol-induced pressor responses dose-dependently. The bradycardic response to carbachol was also inhibited by pirenzepine, but no dose-dependency was observed. Injection of pirenzepine into the basolateral amygdala at a dose (50 nmol) that inhibited carbachol-induced changes in mean arterial pressure and heart rate when injected into the central nucleus of the amygdala failed to exert any inhibition. Methoctramine at a dose of 50 nmol injected into both the central nucleus of the amygdala and the basolateral amygdala did not cause any significant alteration in the responses. These results indicate that muscarinic M1 receptors in the central nucleus of the amygdala are involved in cardiovascular regulation mediated by central cholinergic pathways.

Role of the AV3V region in the pressor responses induced by amygdala stimulation

The role of the anteroventral third ventricle (AV3V) region in the pressor responses to carbachol injected into the lateral cerebral ventricle (i.c.v.), the electrical stimulation of and carbachol-induced stimulation of, the central nucleus of the amygdala were investigated in conscious, unrestrained Sprague-Dawley rats. I.c.v. and intra-amygdalar carbachol caused a significant rise in blood pressure of 22.9 +/- 2.8 and 16.8 +/- 2.2 mmHg, respectively. Electrical stimulation (1 ms, 80 Hz, 50-300 microA, for 30 s) of the central nucleus of amygdala also produced intensity-dependent pressor effects. Electrolytic lesion of the AV3V region abolished the pressor responses induced by carbachol and by electrical amygdala stimulation. The heart rate changes were also significantly inhibited in the AV3V-lesioned rats. These results indicate that the integrity of the AV3V region is essential for the central cholinergic cardiovascular changes induced by central amygdaloid nucleus stimulation.

Effects of combined estrogen and progesterone replacement treatment on detrusor contractility and histology in oophorectomized rats.

Aim:  The aim of this study was to investigate the effects of estrogen plus progesterone treatment administered after surgical menopause on morphological and muscarinic receptor sensitivity in detrusor muscle bands of rats.

Effect of muscimol on cholinomimetic-induced cardiovascular responses in rats

Brain acetylcholine and gamma-aminobutyric acid (GABA) are both involved in the regulation of central cardiovascular control. Despite data from anatomical and electrophysiological experiments characterizing the interaction between central GABAergic and cholinergic neurotransmission, the potential significance of this interaction in central cardiovascular regulation remains unknown. The purpose of this study was to determine whether activation of GABA(A) receptors by intracerebroventricular or intrahypothalamic administration of muscimol affects the cholinergic agonist-induced cardiovascular responses. All experiments were performed in conscious, Sprague-Dawley rats instrumented with a guide cannula for drug injection and iliac arterial catheters for direct measurement of mean arterial pressure and heart rate. Administration of a cholinergic agonist, carbachol, either intracerebroventricularly or into the dorsomedial hypothalamic nucleus, produced a significant increase in mean arterial pressure, whereas injection of carbachol into the posterior hypothalamic nucleus caused a slight elevation in blood pressure. Pretreatment with muscimol 10 min before administration of carbachol prevented the carbachol-evoked blood pressure changes. On the other hand, carbachol produced variable changes in heart rate, depending on the site of injection. In [3H]quinuclydinyl benzilate binding experiments, muscimol did not displace the muscarinic radioligand from its binding sites, suggesting that it does not exert any direct antagonistic activity at muscarinic receptors. These results suggest that the dorsomedial hypothalamic nucleus is a potential site of action for microinjected carbachol and that the GABAergic system has an inhibitory influence on cholinergic neurons involved in blood pressure regulation.

Investigation of the roles of non-neuronal acetylcholine in chronic myeloid leukemic cells and their erythroid or megakaryocytic differentiated lines

BACKGROUND Many studies suggested that Acetylcholine (ACh) might serve as an autocrine/ paracrine growth factor in several types of tumors or tumor cell lines. High levels of Acetylcholinesterase (AChE) activity have been reported in primary brain tumors, ovarian, colon and lung tumors. OBJECTIVES The role of cholinergic signaling needs to be clarified in in leukemia. METHOD K562 cells were derived from a chronic myelogenous leukemia patient during blast crisis serving as pluripotent hematopoietic stem cells. K562 cells were incubated with various cholinergic agonists or antagonists to investigate the role of ACh in different differentiated cell lines. RESULTS Our experiments showed that AChE activity was increased in response to ACh in undifferentiated K562 cells, but in the erythroid differentiated K562 cells a high concentration of ACh (1 mM) decreased the AChE activity. ACh failed to elevate the AChE activity in the megakaryocytic differentiated K562 cells. An AChE inhibitor, eserine, also suppressed the AChE activity in a concentration-dependent manner. Choline uptake inhibition by hemicholinium did increase the AChE activity but not in the erythroid differentiated K562 cell line. Likewise, megakaryocytic differentiated K562 cells also displayed a similar pattern. Vesamicole, a vesicular choline uptake inhibitor, produced similar results. Curare, a nicotinic antagonist, elevated the cell counts of the megakaryocytic differentiated cells. CONCLUSION Our findings may suggest excess extracellular ACh will decrease the cell growth in undifferentiated and megakaryocytic differentiated K562 cell lines through nicotinic type cholinoceptors.