Hideo Negoro

Responses of paraventricular and supraoptic units to angiotensin II, SAR1-ILE8-angiotensin II and hypertonic NaCl administered into the cerebral ventricle

Extracellular recordings of action potentials were made from neurones antidromically identified as neurosecretory cells in the paraventricular and supraoptic nuclei of urethane-anesthetized female rats. Eighty-six neurones were examined for their responsiveness to 10 ng of angiotensin II (AII) injected into the third cerebral ventricle and 78 (91%) of them increased their firing rate following the AII injection. None of the neurosecretory cells tested showed a response to the intraventricular (IVT) injection of isotonic NaCl. Thalamic neurones and non-neurosecretory hypothalamic neurones did not respond to the AII given IVT. Firing activity of 13 neurosecretory neurones was recorded during reflex milk ejection induced by suckling pups in the lactating rats. Seven of them were classified as oxytocinergic cells because they showed a burst of activity before reflex milk ejections and the remaining 6 neurones which gave no burst of firing before milk ejections were classified as nonoxytocinergic neurones. The IVT application of AII resulted in activation of all the oxytocinergic neurones and 5 of the 6 non-oxytocinergic neurones. The effect of AII on the firing of the neurosecretory cell was inhibited by the simultaneous application of Sar1-Ile8-AII (1 microgram), a competitive AII antagonist. The IVT injection of the antagonist alone inhibited the spontaneous firing of the neurosecretory cells, but it did not affect the firing of thalamic or non-neurosecretory hypothalamic neurones. Hypertonic NaCl (0.85 M NaCl, 1 mu1 IVT) also activated 13 of 20 neurosecretory cells tested. Combined application of AII and hypertonic NaCl elicited a marked potentiation of the response of neurosecretory cells to each of the stimuli. These findings indicate that AII activates neurosecretory cells stimulating specific AII receptors in the brain and that AII has a synergistic action with hypertonic NaCl. Inhibition of spontaneous activity of neurosecretory cells by a competitive AII antagonist suggests that endogenous AII may participate in the maintenance of basal activity of neurosecretory cells.

Electrophysiological evidence for multiple sites of actions of angiotensin II for stimulating paraventricular neurosecretory cells in the rat.

Microelectrophoretically (MEPh) applied angiotensin II (AII) excited about half of the PV neurosecretory cells recorded. The excitation was blocked by MEPh applied Sar1-Ala8-AII. Intraventricular (IVT) injection of AII excited both the sensitive cells and insensitive cells to the MEPh applied AII. MEPh-applied Sar1-Ala8-AII, however, blocked the IVT AII induced excitation only in the former type of the cells.

Activation of gastric afferents increases noradrenaline release in the paraventricular nucleus and plasma oxytocin level

Effects of electrical stimulation of the gastric vagal nerves on plasma levels of oxytocin (OXT) and arginine vasopressin (AVP) were examined in rats anesthetized with urethane. Electrical stimulation of the gastric vagal nerves increased the plasma levels of OXT, but not AVP. The concentrations of extracellular noradrenaline (NA) in the paraventricular nucleus (PVN) were measured by in vivo microdialysis in rats anesthetized with urethane. Electrical stimulation of the gastric vagal nerves evoked an increase followed by a slight decrease in the concentrations of NA. The responses of spontaneous firing magnocellular neurosecretory neurons in the PVN to both electrical stimulation of the gastric vagal nerves and intravenous (i.v.) administration of CCK-8 were examined. Most of the putative OXT-secreting cells recorded were excited by both electrical stimulation of gastric vagal nerves and i.v. administration of CCK-8. These results suggest that gastric vagal afferents activate the central noradrenergic system from the brainstem to the PVN and secretion of OXT.

Electrophysiological evidence for neural connections between the supraoptic nuclei.

Effects of electrical stimulation of the contralateral supraoptic nucleus (SON) on the activity of neurosecretory neurons in the SON were studied in urethane-anesthetized lactating rats. Thirty-one out of 41 oxytocin neurons were excited and only two neurons were inhibited by contralateral SON stimulation. Eight out of 26 vasopressin neurons were excited and 8 were inhibited. These responses were not affected by suckling stimuli. Neither oxytocin nor vasopressin neurons tested were antidromically activated by contralateral SON stimulation. Thus monosynaptic connections between the bilateral SON neurosecretory neurons seem to be very few, if any. These results suggest that neural connections exist between the bilateral SON and that they are mainly polysynaptic.

Effects of microelectrophoretically applied acetylcholine- and angiotensin-antagonists on the paraventricular neurosecretory cells excited by osmotic stimuli

Extracellular action potentials were recorded from neurons identified antidromically as neurosecretory cells in the paraventricular nucleus of urethane-anesthetized rats. An intracarotid injection of 0.6 M NaCl increased the firing rate of 26 out of the 33 neurosecretory cells tested. The excitation induced by hypertonic NaCl was blocked by hexamethonium in 6 of 10 neurosecretory cells with nicotinic-cholinergic receptors, by atropine in 4 of 5 cells with muscarinic-cholinergic receptors and by [Sar1, Ala8]-angiotensin II in 5 of 12 cells with angiotensin II receptors. These results suggest that nicotinic- or muscarinic-cholinergic receptors and/or angiotensin II receptors appear to be involved in the final transmission of osmotic stimuli in the paraventricular neurosecretory cells.

Facilitatory effect of antidromic stimulation on milk ejection-related activation of oxytocin neurons during suckling in the rat

Oxytocin neurons of the paraventricular nucleus were identified by their antidromic response to stimulation of the neurohypophysis and a short high-frequency burst of spikes displayed before reflex milk ejection in the urethane-anesthetized lactating rat. During suckling, the milk ejection-related bursts recurred at regular intervals. Stimulation of the neurohypophysis at 50-300 pulses/s for 0.5-6 s (current 1 mA) evoked additional burst(s) with the amplitude higher than those of spontaneously occurring ones. Stimulation with subthreshold current for antidromic response or constant-collision test could also facilitate the neurosecretory bursts. Possibly, oxytocin released within the magnocellular nuclei is responsible for these effects.

Estrogen fails to reduce tuberoinfundibular dopaminergic neuronal activity and to cause a prolactin surge in lactating, ovariectomized rats.

The activity of tuberoinfundibular dopaminergic (TIDA) neurons was estimated by measuring the concentrations of dihydroxyphenylacetic acid (DOPAC) in the median eminence of ovariectomized lactating rats, ovariectomized non-lactating female rats and castrated male rats in correlation with the estrogen-induced prolactin (PRL) surge. PRL surge with a peak at 17.00 h was observed in ovariectomized non-lactating rats but not in the other groups. DOPAC levels were significantly lower at 17.00 h than at 11.00 h in non-lactating female rats, but there was no change of DOPAC levels in lactating rats and male rats. These results indicate that TIDA neuronal activity is inhibited at the time of estrogen-induced PRL surge and that the TIDA neurons may alter their function to result in refractoriness of PRL response to estrogen in the lactating rat.

Control of Oxytocin Secretion in Lactation

The most established physiological function of oxytocin is to induce milk ejection from the mammary gland of lactating animals. It is now known that during lactation oxytocin is released pulsatively following brief periods of burst-like and synchronous activation of many thousands of oxytocin cells in the hypothalamus. The mechanism generating such activity in oxytocin cells has been extensively studied, but it has not been fully understood yet. To explain that suckling stimuli produce a recurrence of milk ejection bursts of oxytocin cells without any change in their background activity, a gating mechanism has been hypothesized. In the excitatory transmission of afferent signals of the milk ejection reflex, alpha adrenergic receptors are indicated to be involved. Among neuropeptides, oxytocin and CRF are potent facilitatory factors. As non-neurochemical factors that facilitate milk ejection bursts of oxytocin cells, there are osmotic stimuli, neurohypophyseal stimulation and vaginal distention. During the lactation period, responsiveness of oxytocin cells to various stimuli such as stress, osmotic stimuli and CCK is markedly reduced. The cause of the change has not been discovered, but it is assumed that the reduction in responsiveness may enable the animal to adapt to the large demands for the hormone during the lactation period.