Minehiro Gotoh

Neostigmine-induced hyperglycemia is mediated by central muscarinic receptor in fed rats

We previously reported that neostigmine injected into the third cerebral ventricle stimulated adrenal secretion of epinephrine, secretion of glucagon from the pancreas, and direct neural innervation of the liver, resulting in hepatic venous plasma hyperglycemia in anesthetized fed rats. However, receptor type of these 3 mechanisms is not known. Therefore, we examined the effects of intraventricularly injected cholinergic or adrenergic antagonists on neostigmine-induced catecholamines in intact rats, glucagon secretion which is mediated by direct neural innervation of pancreas in bilateral adrenalectomized (ADX) rats, and hepatic venous hyperglycemia which is mediated by direct neural innervation of liver in ADX rats receiving constant infusion of somatostatin from femoral vein. Atropine injected into the third cerebral ventricle suppressed epinephrine secretion and dose-dependently inhibited hepatic venous hyperglycemia induced by neostigmine in intact rats. The neostigmine-induced glucagon secretion which occurs in ADX rats was suppressed by atropine. Atropine also prevented the neostigmine-induced hyperglycemia in ADX rats receiving constant somatostatin infusion through femoral vein (ADX-Somato rats). On the other hand, phentolamine, propranolol and hexamethonium showed no significant inhibitory effect on neostigmine-induced hyperglycemia, epinephrine and glucagon secretion in intact rats, glucagon secretion in ADX rats, or hyperglycemia in ADX-Somato rats. These results suggest that neostigmine-induced epinephrine and glucagon secretion and increased hepatic glucose output stimulated by direct neural innervation to liver is mediated by central muscarinic receptor in fed rats.

Swimming stress that causes hyperglycemia increases in vivo release of noradrenaline, but not acetylcholine, from the hypothalamus of conscious rats.

The effects of acute swimming stress (10 min) on noradrenaline release from the medial basal hypothalamus (MBH; consisting of the ventromedial and dorsomedial hypothalamus) and acetylcholine release from the lateral hypothalamic area (LHA) were investigated in freely moving rats by using in vivo microdialysis techniques. Serum glucose, noradrenaline and adrenaline concentrations were also determined. Acute swimming stress produced significant hyperglycemia, with increases in serum noradrenaline and adrenaline concentrations. The release of noradrenaline from the MBH was significantly stimulated during the swimming stress. On the other hand, the swimming stress has no significant effect on the release of acetylcholine from the LHA. These findings support the idea that hypothalamic noradrenergic neurons play an important role in the sympathoadrenal hyperglycemic response to stressful stimuli. Moreover, it is suggested that hypothalamic cholinergic neurons are not involved in the responses of serum glucose, noradrenaline and adrenaline concentrations to swimming stress.

Vagally mediated insulin secretion by stimulation of brain cholinergic neurons with neostigmine in bilateral adrenalectomized rats

This study investigated the relationship between central cholinergic neurons and insulin secretion in bilateral adrenalectomized fed rats. Neostigmine (a cholinesterase inhibitor, 5 x 10(-8) mol) administered into the third cerebral ventricle produced significant increases in hepatic venous plasma insulin and glucose concentrations, whereas i.v. injection of the same dose of neostigmine did not. Prior acute subdiaphragmatic vagotomy or i.p. pre-injection with methylatropine (10(-8) mol) completely prevented the neostigmine-induced rise in plasma insulin concentration. Intraperitoneal pretreatment with hexamethonium (5 x 10(-8) mol) also significantly reduced the plasma insulin response. These peripheral pretreatments did not change the plasma glucose response to neostigmine. Intraventricular co-administration of 10(-9) mol methylatropine, a dose that was ineffective when pre-injected i.p., eliminated the plasma insulin and glucose responses to neostigmine, whereas hexamethonium (5 x 10(-8) mol) had no influence on either response to neostigmine. These observations suggest that stimulation of central cholinergic-muscarinic neurons with third cerebral ventricular injection of neostigmine results in vagally mediated insulin secretion in bilateral adrenalectomized fed rats.

Hyperglycemia induced by hippocampal administration of neostigmine is suppressed by intrahypothalamic atropine

We investigated the relationship between the hyperglycemia induced by the administration of neostigmine into the hippocampus and the hypothalamus. Prior to the injection of neostigmine (5 x 10(-8) mol) into the hippocampus, 1 microliter each of atropine or hexamethonium (5 x 10(-11)-5 x 10(-8) mol) was injected into the bilateral ventromedial hypothalamus (VMH). Atropine suppressed in a dose-dependent manner the hyperglycemia induced by hippocampal administration of neostigmine, whereas hexamethonium had no significant effect. These observations suggest that the pathway for this experimental hyperglycemia involves, at least in part, the muscarinic cholinergic neurons in the VMH.

Effects of intracerebroventricularly administered neostigmine on hypothalamic monoaminergic neuronal activities in awake rats

Hypothalamic neuronal activities of noradrenaline, dopamine and serotonin as well as serum glucose concentrations were simultaneously monitored 60 min after the third cerebroventricular injection of neostigmine, a cholinesterase inhibitor, in rats. Each neuronal activity was assessed from a ratio of the concentration of the major metabolite to that of its precursor monoamine itself by using the technique of gas chromatography-mass spectrometry. Neostigmine caused significant increases in serum glucose concentrations, hypothalamic noradrenergic and dopaminergic neuronal activities, and significantly suppressed hypothalamic serotonergic neuronal activity. All these responses to neostigmine were completely inhibited by the co-administration of atropine. These observations emphasize the important role of the interactions between cholinergic (muscarinic) and monoaminergic neurons in the brain.

Effects of intracerebroventricularly administered neostigmine on sympathetic neural activities of peripheral tissues in rats

Sympathetic nervous activity (SNA) in the liver, heart, pancreas and interscapular brown adipose tissue was examined 60 min after the third cerebroventricular injection of neostigmine (5 x 10(-8) mol) in rats. We employed the technique of specific gas chromatography-mass spectrometry for simultaneous analysis of norepinephrine (NE) and its primary neuronal metabolite, 3,4-dihydroxyphenylethyleneglycol (DHPG) and used the ratio DHPG/NE as an index of SNA. Neostigmine produced significant increases in the DHPG/NE ratio in all tissues investigated. Co-administration of atropine with neostigmine completely inhibited this neostigmine-induced effect. These findings suggest that the central cholinergic-muscarinic activation with neostigmine stimulates SNA in the peripheral tissues examined.

Central Versus Peripheral Effect of Clonidine on Hepatic Venous Plasma Glucose Concentrations in Fasted Rats

To evaluate whether clonidine exerts its action within the central nervous system or outside the central nervous system to induce hyperglycemia, we compared the effects of clonidine injected into the third cerebral ventricle or intravenously on hepatic venous plasma glucose concentrations in fasted rats. Clonidine administration produced a dose-dependent hyperglycemia in each case. At all tested doses (5, 50, and 100 nmol) the hyperglycemic responses to clonidine injected intravenously were not significantly different from those to clonidine injected into the third cerebral ventricle. Intraperitoneal pretreatment with yohimbine (50 nmol) or phentolamine (50 nmol), both α-adrenergic antagonists, reduced the hyperglycemic response to clonidine (100 nmol) given intravenously, but these antagonists preinjected into the third cerebral ventricle did not reduce the response. Moreover, no significant differences in venous plasma clonidine concentrations were observed when intravenous and third cerebral ventricle injections of clonidine (100 nmol) were compared. These results suggest that clonidine-induced hyperglycemia is mainly mediated by the peripheral mechanism rather than through the central mechanism. To gain an insight into the peripheral mechanism for the hyperglycemic action of clonidine, we measured the plasma immunoreactive glucagon and insulin concentrations after intravenous clonidine (100 nmol). We found that plasma immunoreactive glucagon concentrations significantly increased, whereas plasma immunoreactive insulin concentrations did not change significantly despite hyperglycemia. Furthermore, the clonidine-induced hyperglycemia and hyperglucagonemia were completely abolished by constant intravenous infusion of somatostatin (1.0 μg. kg− · min−1). These results suggest that clonidine-induced hyperglycemia is associated with not only the suppression of insulin release but also the stimulation of glucagon release. Additionally, clonidine may not act directly on the liver to produce hyperglycemia in fasted rats.

Hypothalamic monoamine metabolism is different between the diabetic GK (Goto-Kakizaki) rats and streptozotocin-induced diabetic rats.

Hypothalamic neuronal activities of noradrenaline, dopamine and serotonin were investigated in the GK (Goto-Kakizaki) rats, a model for type 2 diabetes, and the streptozotocin (STZ)-induced diabetic rats, a model for type 1 diabetes. Each neuronal activity was assessed from a ratio of the concentrations of the major metabolite to those of its precursor monoamine itself or the metabolite concentrations itself. In the GK rats, hypothalamic noradrenaline and dopamine activities were significantly increased, and serotonin activity was unchanged. In the STZ-induced diabetic rats, hypothalamic dopamine and serotonin activities were significantly decreased, and noradrenaline activity was unchanged. In these two diabetic rats, hypothalamic noradrenaline and dopamine activities correlated significantly with serum insulin concentrations, whereas hypothalamic serotonin activity correlated significantly with serum glucose concentrations. These findings suggest that hypothalamic monoamine metabolism of the GK rats is quite different from that of the STZ-induced diabetic rats, and that not only hyperglycemia but also serum insulin concentrations have an influence on the hypothalamic monoamine metabolism in these diabetic rats.

Octreotide-induced suppression of the hyperglycemic response to neostigmine or bombesin: relationship to hypothalamic noradrenergic drive

Neostigmine (cholinesterase inhibitor) or bombesin, when injected into the third cerebral ventricle of awake rat, dose-dependently increased serum glucose with the simultaneous rise in hypothalamic noradrenergic neuronal activity (NAA). Co-administration of octreotide with neostigmine or bombesin suppressed the hypothalamic NNA response with the simultaneous inhibition of the hyperglycemic response. There was a close relationship between hypothalamic NNA and serum glucose in these studies. On the basis of the concept that hypothalamic noradrenergic drive plays an important role in mediating the hyperglycemic response to stressful stimuli, the present findings suggest that the hyperglycemic response to neostigmine or bombesin is mediated via the interaction with hypothalamic noradrenergic neurons.

Starvation reduces norepinephrine activities in both hypothalamus and heart in rats

Norepinephrine (NE) activities in both hypothalamus and heart were simultaneously assessed in rats after food-deprivation for 2 days. The technique of gas chromatography-mass spectrometry was employed for the analysis of NE and its primary neuronal metabolite, 3,4-dehydroxyphenylethylene glycol (DHPG), and the ratio of DHPG to NE was used as an index of NE activity. Hypothalamic DHPG/NE ratio was significantly decreased by fasting and was completely reversed by a single day of refeeding. These changes in hypothalamic DHPG/NE ratio were parallel to those in cardiac DHPG/NE ratio. Supporting the concept in which hypothalamic NE neurons play an important role in modulating the sympathetic outflow, it is suggested the decrease in hypothalamic NE activity contributes to the reduction in cardiac NE activity during fasting.