Hiroko Miyakubo

Involvement of serotonergic systems in the lateral parabrachial nucleus in sodium and water intake: a microdialysis study in the rat.

The present study was carried out to investigate whether 0.3 M NaCl and water intake alters the release of serotonin (5-hydoxytryptamine, 5-HT) in the region of the lateral parabrachial nucleus (LPBN) in freely moving rats. The ingestion of 0.3 M NaCl and water was induced by subcutaneous injections of the diuretic furosemide (FURO, 10 mg/kg) and the angiotensin converting enzyme inhibitor captopril (CAP, 5 mg/kg), and extracellular concentrations of 5-HT and its metabolite 5-hydroxyindoleacetic acid (5-HIAA) were measured using intracerebral microdialysis techniques. The intake of 0.3 M NaCl and water significantly enhanced the 5-HT and 5-HIAA levels in the LPBN area. The combined treatment with FURO and CAP elicited significant decreases in the 5-HT and 5-HIAA concentrations in the LPBN area under the condition that 0.3 M NaCl and water are not available for drinking. These results suggest that the serotonergic system in the LPBN area may play an important role in the modulation of sodium appetite and thirst.

Angiotensinergic and noradrenergic mechanisms in the hypothalamic paraventricular nucleus participate in the drinking response induced by activation of the subfornical organ in rats

The present study was done to investigate the contribution of the hypothalamic paraventricular nucleus (PVN) to the drinking response caused by activation of the subfornical organ (SFO) following angiotensin II (ANG II) injections in the awake rat. Microinjection of ANG II into the SFO elicited the drinking response. Previous injections of either saralasin, an ANG II antagonist, or phentolamine, an alpha-adrenoceptor antagonist, bilaterally into the PVN resulted in the significant attenuation of the drinking response to ANG II. Similar injections of any of the beta-adrenoceptor antagonist timolol, the muscarinic antagonist atropine, or saline vehicle into the PVN had no significant effect on the drinking response. In an attempt to clarify the neural mechanisms in the PVN involved in the drinking response to ANG II injected into the SFO, the effect of microinjection of ANG II into the SFO on noradrenaline (NA) release in the PVN was examined using intracerebral microdialysis techniques. The injection of the ANG II, but not saline vehicle, significantly enhanced the NA release in the region of the PVN. These results indicate the involvement of both the angiotensinergic and alpha-adrenergic systems in the PVN in the drinking response caused by angiotensinergic activation of the SFO, and imply that the angiotensinergic projections from the SFO to the PVN may serve to increase NA release which results in mediating water intake.

Estrogen decreases the responsiveness of subfornical organ neurons projecting to the hypothalamic paraventricular nucleus to angiotensin II in female rats

Extracellular single-unit activity was recorded from subfornical organ (SFO) neurons antidromically identified as projecting to the hypothalamic paraventricular nucleus (PVN) in urethane-anesthetized ovariectomized female rats that were treated with either propylene glycol (PG) vehicle or estradiol benzoate (EB). No significant differences were observed between the PG- and EB-treated rats in the latency, conduction velocity, or threshold of antidromic activation. The mean spontaneous firing rate was significantly lower and the refractory period was significantly longer in the EB-treated rats. In the identified units that were activated by angiotensin II (ANG II) applied iontophoretically, the amount of excitatory response to intracarotid administration of ANG II was much greater in the PG-treated than in the EB-treated rats. These results suggest that estrogen may decrease the responsiveness of SFO neurons projecting to the PVN to circulating ANG II.

Enhanced response of subfornical organ neurons projecting to the hypothalamic paraventricular nucleus to angiotensin II in spontaneously hypertensive rats

Thirty subfornical organ (SFO) neurons in normotensive Wistar-Kyoto (WKY) rats and 32 SFO neurons in spontaneously hypertensive rats (SHR) were antidromically activated by electrical stimulation of the hypothalamic paraventricular nucleus (PVN) under urethane anesthesia. The spontaneous firing rate was significantly higher in SHR than in WKY rats. No significant differences in the latency, conduction velocity, and threshold of antidromic response were observed between WKY and SHR. All the identified SFO units were tested for a response to intracarotid injection of angiotensin II (ANG II, 20-ng/kg b.w.t.). Injections of ANG II elicited an increase in the activity of 21 units in WKY and 20 units in SHR and a depression in the firing of one unit in WKY rats, but did not affect the remaining units. The magnitude of the excitatory response caused by the ANG II injection was much greater in SHR than in WKY rats. These results show that there are differences between WKY and SHR in the spontaneous discharge rate of SFO neurons projecting to the PVN and in their response to circulating ANG II.

Activation of serotonergic pathways from the midbrain raphe system to the subfornical organ by hemorrhage in the rat.

The role of serotonergic neural pathways from the midbrain raphe nuclei to the subfornical organ (SFO) in the central regulation of cardiovascular function and body fluid balance was investigated in adult male rats under urethane anesthesia. Eleven neurons in the dorsal raphe nucleus (DR) were antidromically activated by electrical stimulation of the SFO. Of these neurons, 6 displayed an excitatory response following hemorrhage (10 ml/kg bwt) while the remaining 5 neurons were unresponsive. Ninety-four neurons in the SFO were tested for a response to electrical stimulation of the DR or hemorrhage. Electrical stimulation of the DR caused orthodromic excitation (19%) or inhibition (5%) of the activity of SFO neurons. In 14 of 18 SFO neurons that displayed the excitation to the stimulation of the DR, hemorrhage (30 to 50 mm Hg suppression in mean arterial pressure) produced an increase of their discharge, while the stimulus was without effect in the remaining neurons responsive to the stimulation of the DR. The effects of hemorrhage on serotonin (5-HT) release in the region of the SFO were examined using intracerebral microdialysis techniques. Hemorrhage significantly increased 5-HT and its metabolite 5-hydroxyindoleacetic acid (5-HIAA) concentrations in the region of the SFO. The present data suggest that the serotonergic pathways from the DR to the SFO may relay activation of the peripheral baroreceptors to SFO neurons which result in enhanced excitability, indicating the involvement of the pathways in the regulation of cardiovascular function.

Reduced Dipsogenic Response Induced by Angiotensin II Activation of Subfornical Organ Projections to the Median Preoptic Nucleus in Estrogen-Treated Rats

The present study was carried out to investigate whether estrogen modulates the drinking response induced by activation of angiotensinergic neural pathways from the subfornical organ (SFO) to the median preoptic nucleus (MnPO). Microinjection of angiotensin II (ANG II, 10(-10) M, 0.2 microl) into the SFO elicited drinking in ovariectomized (OVX) female rats that were treated with either propylene glycol (PG) vehicle or estrogen benzoate (EB). The amount of water intake induced by the ANG II injection was significantly greater in the PG-treated than in the EB-treated animals. In both groups of female rats, previous injections of saralasin (Sar, 10(-10) M, 0.2 microl), a specific ANG II antagonist, into the MnPO resulted in the significant attenuation of the drinking response to ANG II, showing that the ANG II-induced drinking response may be mediated in part by the angiotensinergic SFO projections to the MnPO. Injections of ANG II (10(-10) M, 0.2 microl) into the MnPO caused drinking in both groups, while no significant difference was found between the groups in the amount of water intake. These results suggest that increases in the circulating level of estrogen may attenuate the drinking response induced by ANG II activation of the SFO projections to the MnPO.

GABAergic modulation of noradrenaline release in the median preoptic nucleus area in the rat.

Microdialysis was employed to investigate whether gamma-aminobutyric acid (GABA) receptor mechanisms are involved in the regulation of noradrenaline (NA) release in the median preoptic nucleus (MnPO) in awake, freely moving rats. Perfusion with the GABA receptor antagonists as well as agonists was performed in the region of the MnPO through a microdialysis probe and dialysate levels of NA were measured. Perfusion with either bicuculline (10 and 50 microM), a GABA(A) receptor antagonist, or phaclofen (10 and 50 microM), a GABA(B) receptor antagonist, enhanced the release of NA in the MnPO area. Higher-dose perfusion with the GABA(A) agonist muscimol (50 microM) or the GABA(B) agonist baclofen (250 microM) decreased dialysate NA in the MnPO area. An iso-osmotic reduction of fluid volume following subcutaneous treatment with polyethylene glycol (PEG, 30%, 5 ml) significantly increased the NA level in the MnPO area. The increased levels of NA caused by the PEG treatment were attenuated by perfusion with muscimol (10 microM), but not by baclofen (50 microM). These results show the participation of both GABA(A) and GABA(B) receptors in the modulation of the release of NA in the MnPO area, and imply that the GABA(A) receptor mechanism may play an important role in the noradrenergic regulatory system of body fluid balance.

Noradrenaline release in the median preoptic nucleus area caused by hemorrhage in the rat

The present study was designed to examine whether noradrenergic projections from the A1 cell group in the ventrolateral medulla to the median preoptic nucleus (MnPO) transmit information from the peripheral baroreceptors. In urethane-anesthetized male rats, extracellular concentrations of noradrenaline (NA) in the region of the MnPO in response to hemorrhage (5 or 10 ml/kg) were monitored with in vivo microdialysis methods. Hemorrhage significantly increased the NA release in the MnPO area. The enhancement of NA release in the MnPO area caused by hemorrhage was significantly attenuated by previous injections of the local anesthetic lidocaine (2 %, 0.2 microl), but not by saline (0.2 microl), into the A1 region. These results suggest that the noradrenergic projections from the A1 region are important for carrying the peripheral baroreceptor information to the MnPO.

Attenuated drinking response induced by angiotensinergic activation of subfornical organ projections to the paraventricular nucleus in estrogen-treated rats

The present study was carried out to examine whether estrogen modulates the drinking response caused by activation of neural pathways from the subfornical organ (SFO) to the hypothalamic paraventricular nucleus (PVN) in the female rat. Microinjection of angiotensin II (ANG II) into the SFO elicited drinking in ovariectomized female rats that were treated with either propylene glycol (PG) vehicle or estradiol benzoate (EB). The amount of water intake induced by the ANG II injection was significantly greater in the PG-treated than in the EB-treated animals. In both groups, previous injections of either saralasin, an ANG II antagonist, or phentolamine, an alpha-adrenoceptor antagonist, bilaterally into the PVN resulted in the significant attenuation of the drinking response to ANG II, whereas similar injections of saline vehicle into the PVN were without effect. These results suggest that the circulating estrogen may act to reduce the drinking response that is mediated through angiotensinergic and alpha-adrenergic mechanisms in the PVN in response to angiotensinergic activation of SFO efferent projections.

GABAergic modulation of neurons in the nucleus of the solitary tract with ascending projections to the subfornical organ in the rat

Twenty-five neurons in the region of the nucleus of the solitary tract (NTS) were antidromically activated by electrical stimulation of the subfornical organ (SFO) in male rats under urethane anesthesia. Microiontophoretically applied bicuculline, a gamma-aminobutyric acid (GABA)(A) antagonist, but not phaclofen, a GABA(B) antagonist, attenuated the post-antidromic inhibitory response evoked by SFO stimulation of approximately two-third (n=17) of identified neurons, indicating the existence of recurrent inhibitory systems through GABA(A) receptors. Iontophoretically applied GABA decreased the spontaneous activity of all identified neurons, and the GABA-induced inhibition was prevented by simultaneously applied bicuculline, but not by phaclofen. Activation of peripheral baroreceptors, achieved by rising arterial blood pressure with an intravenous infusions of phenylepherine, suppressed the activity of the majority (n=20) of identified neurons. The inhibitory response of identified neurons (n=7) to baroreceptor activation was partially antagonized by iontophoretically applied bicuculline, but not by phaclofen. These results imply that GABAergic mechanisms may modulate the baroreceptor reflex acting on GABA(A) receptors of NTS neurons with ascending projections to the SFO in the region of the NTS.