Cell subpopulations of nicotine-sensitive subfornical organ neurons in rat

The subfornical organ (SFO), which is related to drinking and cardiovascular regulation, is activated by central application of nicotine (NIC) and angiotensin II (ANG). However, NIC-induced drinking is much smaller than ANG-induced one although approximately 60% of SFO neurons are affected by both NIC and ANG. Therefore, some specific subpopulations of SFO neurons for NIC or ANG may be related to such different drinking responses. To clarify subpopulations of NIC-sensitive neurons, electrophysiological properties of SFO neurons with the application of NIC was investigated at whole-cell patch-clamp recordings. Based on our developed electrophysiological criteria of the recovery kinetics of tetraethylammonium-resistant transient outward K(+) currents, two sub-types (F- and S-type neurons) were distinguished. Twenty-nine dissociated SFO neurons were examined to determine whether they showed NIC-induced inward currents. Most F-type neurons (n=19/21) showed NIC sensitivity, but most S-type neurons (n=7/8) did not. Our previous study had demonstrated that half of the F-type and all of the S-type units showed ANG sensitivity. These suggests that almost all of the NIC-sensitive SFO neurons were electrophysiologically classified as the F-type, but not S-type, and this differs in part from angiotensin sensitivity. The different subpopulations for chemical sensitivities in the SFO may be related to different drinking responses.     

Region-specific projections from the subfornical organ to the paraventricular hypothalamic nucleus in the rat

Neurons in the subfornical organ (SFO) project to the paraventricular hypothalamic nucleus (PVN) and there, in response to osmolar and blood pressure changes, regulate vasopressin neurons in the magnocellular part (mPVN) or neurons in the parvocellular part (pPVN) projecting to the cardiovascular center. The SFO is functionally classified in two parts, the dorsolateral peripheral (pSFO) and ventromedial core parts (cSFO). We investigated the possibility that neurons in each part of the SFO project region-specifically to each part of the PVN, using anterograde and retrograde tracing methods. Following injection of an anterograde tracer, biotinylated dextran amine (BDX) in the SFO, the respective numbers of BDX-uptake neurons in the pSFO and cSFO were counted and the ratio of the former to the latter was obtained. In addition, the respective areas occupied by BDX-labeled axons per unit area of the mPVN and pPVN were measured and the ratio of the former to the latter was obtained. Similarly, following injection of the retrograde tracer in the PVN, the respective areas occupied by tracer per unit area of the mPVN and pPVN were measured and the ratio of the former to the latter was obtained. The respective numbers of retrogradely labeled neurons in the pSFO and cSFO were also counted and the ratio of the former to the latter was obtained. It became clear by statistical analyses that there are strong positive correlations between the ratio of BDX-uptake neuron number in the SFO and the ratio of BDX-axon area in the PVN in anterograde experiment (correlation coefficient: 0.787) and between the ratio of retrograde neuron number in the SFO and the ratio of tracer area in the PVN in retrograde experiment (correlation coefficient: 0.929). The result suggests that the SFO projects region-specifically to the PVN, the pSFO to the mPVN and the cSFO to the pPVN.     

外周给予内毒素后穹窿下器细胞凋亡的时程变化

目的:研究穹窿下器细胞在外周给予内毒素后的凋亡变化。材料和方法:实验动物用SD大鼠,应用扫描电镜和TUNEL检测方法,分别观察腹腔注射大肠杆菌内毒素2、6、8和16小时后穹窿下器的形态学改变。结果:穹窿下器处发现有TUNEL阳性细胞,扫描电镜观察室管膜细胞及膜上结构随着时间的变化而形态各异。结论:外周给予内毒素引起穹窿下器细胞调亡,而且作为感受性室周器官较其他脑区更早地发生了凋亡,提示穹窿下器很可能是优先感受血携免疫信息的窗口之一。     

杂色曲霉素对小鼠穹窿下器室管膜细胞超微结构的影响

目的：观察杂色曲霉素（ST）对小鼠穹隆下器（SFO）室管膜细胞超微结构的影响。方法：BALB／c小鼠,单次ST（3000μg/kg）灌胃,分别于灌胃后1、2、4、8、16h处死动物,用扫描电镜观察SFO室管膜的结构变化。结果：ST灌胃后1h部分SFO室管膜表面微绒毛结构消失、细胞破损,ST灌胃后2h室管膜出现凹凸不平、损伤加重。ST灌胃后8h室管膜凹凸不平和破损最为明显,ST灌胃后16h逐渐恢复。结论：提示ST对小鼠SFO室管膜细胞有明显的损伤作用。     

实验性变态反应性脑脊髓炎大鼠穹窿下器室管膜细胞的形态学改变

目的：观察穹隆下器(SFO)室管膜细胞在实验性变态反应性脑脊髓炎(EAE)时的形态学变化，探讨其病变的意义。方法：建立Wistar大鼠EAE模型，应用光镜、扫描电镜、透射电镜方法，动态观察EAE发病的潜伏期、发病期、恢复期SFO室管膜细胞的形态学改变。结果：(1)SFO室管膜细胞病变早于EAE发病，且与EAE病情明显正相关。(2)恢复期室管膜下层出现密集排列的染色质浓染的新生细胞。结论：(1)SFO参与了EAE时脑内的免疫反应；而且是EAE发病的早期病变之一，可能是外周免疫介质优先入脑位点。(2)室管膜下神经干细胞可被激活，对室管膜进行修复。     

伽玛刀照射后胶质瘤大鼠穹隆下器的扫描电镜观察

目的：探讨伽玛刀照射大鼠脑胶质瘤前后，穹隆下器(subfornical organ，SFO)室管膜面发生的微观变化。方法：建立C6大鼠脑胶质瘤模型，以胶质瘤为靶区对大鼠行伽玛刀照射，用扫描电镜观察伽玛刀照射前后SFO的形态学变化。结果：在胶质瘤生长期，SFO室管膜细胞出现胞体固缩、胞膜凹陷，细胞表面微绒毛、微突起和单根纤毛减少，室管膜面纤维网发生缠结、断裂；伽玛刀照射后上述变化进一步加重，除胞膜凹陷外，SFO室管膜细胞还出现胞膜破裂，胞核裸露，室管膜面纤维网完全脱落。结论：在胶质瘤生长和伽玛刀照射后肿瘤坏死过程中，SFO室管膜细胞的形态出现明显变化，可能与SFO参与神经免疫调节机制有关。     

Galanin of the homologous species inhibits insulin secretion in the rat and in the pig

It is known that pig galanin inhibits insulin secretion in dogs, rats and mice. The present study examined whether species-specific, homologous, galanin inhibits insulin secretion. Thus, the effects of rat galanin were examined in the rat, and the effects of pig galanin were examined in the pig, both in vivo and in vitro. In conscious rats, synthetic rat galanin (2 nmol kg^-1) abolished the glucose- (0.56 mmol kg^-1) induced increase in plasma insulin levels. In vitro, rat galanin (10^-9 to 10^-6 mol l^-1) inhibited glucose- (8.3 mmol l^-1) stimulated insulin release from isolated rat islets. In anaesthetized pigs, 15 min infusion of synthetic pig galanin (207 pmol min^-1) into the pancreatic artery decreased the insulin output with a subsequent recovery. In vitro, pig galanin (10^-6 mol l^-1) inhibited glucose- (8.3 mmol l^-1) stimulated insulin release from isolated pig islets. We conclude that homologous galanin inhibits insulin secretion in both the rat and the pig.     

甘丙肽抑制大鼠垂体腺瘤细胞体外侵袭性

目的 探讨甘丙肽对大鼠垂体腺瘤细胞侵袭性的作用及其受体机制.方法 提取大鼠垂体腺瘤GH3细胞RNA,反转录后测定甘丙肽及其3个受体亚型的表达情况;将大鼠垂体腺瘤GH3细胞分为对照组、甘丙肽药物处理组和选择性甘丙肽2型受体激动剂AR-M1896组,利用MTT方法检测对照组和实验组在给药后12、24和36 h各分组细胞活力...     

Subfornical organ efferents enhance extracellular noradrenaline concentrations in the median preoptic area in rats

The present study was carried out to examine whether angiotensinergic pathways from the subfornical organ (SFO) regulate the noradrenergic system in the median preoptic nucleus (MnPO). Intracerebral microdialysis techniques were used to quantify the extracellular concentration of noradrenaline (NA) in the MnPO area. In urethane-anesthetized male rats, electrical stimulation (5–20 Hz, 0.6 mA) of the SFO significantly increased the NA concentration in the MnPO area, and the increase was significantly diminished by pretreatment with the angiotensin II (ANG II) antagonist saralasin (Sar; 5 μg), into the third ventricle (3V). Injections of ANG II (5 μg) into the 3V significantly enhanced NA release in the MnPO area. The data imply that the angiotensinergic pathways from the SFO to the MnPO may act to enhance NA release in the MnPO area.     

Diversity of the muscarinic and nicotinic responses of subfornical organ neurons in rat slice preparations

Recently we found that subfornical organ (SFO) neurons were activated through nicotinic receptors as well as muscarinc. In this study, the preferential responses of single SFO neurons to both muscarine and nicotine were examined by using rat slice preparations, and current and voltage clamp recordings. When the amplitudes of the depolarizations and inward currents by muscarine and nicotine in single SFO neurons were compared, some neurons showed a higher sensitivity to muscarine than to nicotine while others showed vice versa. These data indicate that acetylcholine activates SFO neurons preferentially through either muscarinic or nicotinic receptors and suggest a diversity of cholinergic responses in the SFO.     

Activation of metabotropic glutamate receptors inhibits GABAergic transmission in the rat subfornical organ

Glutamate is known to increase neuronal excitability in the subfornical organ, a circumventricular organ devoid of the blood-brain barrier. To understand the synaptic mechanism of neuronal excitation by glutamate in this nucleus, we examined the effects of glutamate on GABAergic spontaneous inhibitory postsynaptic currents recorded from subfornical organ neurons in the rat brain slice. The baseline frequency, amplitude and decay time-constant of such spontaneous synaptic currents were 5.60 Hz, 119 pA and 17.3 ms, respectively. Glutamate (10-1000 microM) selectively inhibited the frequency of spontaneous GABAergic inhibitory postsynaptic currents (half-maximal effective concentration=47 microM) with little effects on their amplitudes and decay time constants. The inhibitory effect of glutamate on the frequency of spontaneous GABAergic postsynaptic currents was not blocked by tetrodotoxin (1 microM), or by the antagonists of ionotropic glutamate receptors. In contrast, such inhibitory effect of glutamate was mimicked by general or group II selective metabotropic glutamate receptor agonists such as DCGIV (2S,1'R,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine (half-maximal effective concentration=112 nM), but not by the agonists for group I or group III metabotropic glutamate receptors. Under current clamp mode, glutamate reduced the frequencies of spontaneous inhibitory postsynaptic potentials and action potentials in subfornical organ neurons. Our data indicate that glutamate decreases the frequency of spontaneous inhibitory postsynaptic currents by acting on the group II metabotropic glutamate receptors on axonal terminals in the subfornical organ. From these results we suggest that the glutamate-induced modulation of tonic GABAergic inhibitory synaptic activity can influence the excitability of subfornical organ neurons.     

Thirst and vasopressin secretion following central administration of angiotensin II in rats with lesions of the septal area and subfornical organ

Various dipsogenic stimuli, including peripheral and central administration of angiotensin II, have been shown to be capable of releasing vasopressin from the neurohypophyseal system. Studies were carried out in the rat to investigate whether the septal area, which contains a high concentration of angiotensin-sensitive cells and has neural connections with hypothalamic vasopressin-secreting neurons, mediated the stimulatory effect produced by angiotensin II on vasopressin release. Rats with electrolytic lesions in the region of the septal area had increased daily water consumption and urine output when these lesions included the medioventral or lateral nuclei of the septal forebrain, but not when the lesion involved the subfornical organ. No difference was observed in drinking responses following water deprivation or intracerebroventricular injection of angiotensin II in all experimental groups. In addition, the impaired ability to maintain water homeostasis (polyuro-polydipsic syndrome) of septal-lesioned rats was associated septal-lesioned rats was associated with decreased levels of circulating radioimmunoassayable vasopressin. Furthermore, the vasopressin release which occurred in response to intracerebroventricular angiotensin II in normal controls, sham-lesioned and subfornical organ-lesioned rats was significantly attenuated in rats with electrolytic lesion of the medioventral or lateral septal area. Since cells in the lateral septal area are excited by iontophoretic application of angiotensin II, the present data might be consistent with the hypothesis that the stimulatory effect produced by central administration of angiotensin II on vasopressin release rests upon the integrity of the lateral septal area.     

Acute electrical stimulation of the subfornical organ induces feeding in satiated rats

The SFO, a circumventricular organ (CVO) that lacks the normal blood-brain barrier, is an important site in central autonomic regulation. A role for the SFO in sensing circulating satiety signals has been suggested by electrophysiological studies demonstrating that the anorexigenic satiety signals, leptin and amylin, as well as the orexigenic satiety signal, ghrelin, influence the excitability of separate populations of SFO neurons. The present study examined whether acute, short duration, electrical stimulation of the SFO influenced feeding in satiated rats. Electrical stimulation (200 µA) of satiated animals with subfornical organ (SFO) electrode placement (n = 6) elicited feeding in all animals tested with a mean latency to eat of 8.0 ± 4.0 min after termination of SFO stimulation (mean food consumption: 0.6 ± 0.12 g/100 g bw). These same rats undergoing a sham stimulation did not eat (mean food consumption: 0.0 ± 0.0 g, n = 6) nor did animals receiving stimulation with non-SFO electrode placements (mean food consumption: 0.0 ± 0.0 g, n = 6). SFO stimulation at this intensity elicited drinking in 5/6 animals with a mean latency to drink of 15.2 ± 2.6 min. Feeding effects were specific to higher stimulation intensities as lower intensity stimulation (100 µA, n = 6) elicited drinking (mean latency to drink: 6.2 ± 2.6 min) but did not cause any animal to eat.The results of the present study show that acute, short duration, SFO stimulation induces feeding in satiated rats, lending support for a role for the SFO as an integrator of circulating peptides that control feeding.     

Efferent pathways from the region of the subfornical organ to hypothalamic paraventricular nucleus: an electrophysiological study in the rat

Summary Twenty-three neurons in the region of the subfornical organ (SFO) were antidromically activated by electrical stimulation of the hypothalamic paraventricular nucleus (PVN) in male rats under urethane anesthesia. Microiontophoretically (MIPh) applied angiotensin II (AII) excited the activity of all units in the region of the SFO and the effect of AII was blocked by MIPh applied saralasin (Sar), an AII antagonist, but not by atropine (Atr), a muscarinic antagonist. In these units, 12 were also excited by MIPh applied acetylcholine (ACh) while 11 were not affected and the effect of ACh was attenuated by not only MIPh applied Atr, but also Sar, suggesting that not only neurons specific for AII, but also neurons sensitive to both AII and ACh project to the PVN in the region of the SFO. Intravenously administered AII excited the activity of both types of units in the region of the SFO. Microinjected AII or ACh into the region of the SFO excited the activity of putative vasopressin (VP)-secreting units in the PVN. These results suggest that neurons projecting to the PVN in the region of the SFO may act to enhance the activity of putative VP-secreting neurons in the PVN in response to circulating AII.     

Involvement of the median preoptic nucleus in the regulation of paraventricular vasopressin neurons by the subfornical organ in the rat

Summary Extracellular recordings in urethane-anesthetized male rats indicated that electrical stimulation of the subfornical organ (SFO) alters the activity of 54 out of 62 phasically firing neurosecretory neurons in the hypothalamic paraventricular nucleus (PVN); 44 cells demonstrate an increase in excitability; 10 cells display a depression in their activity. In 14 out of 38 PVN cells tested, SFO stimulation-evoked excitations were abolished by pretreatment with the angiotensin II (ANG II) antagonist, saralasin (Sar), in the region of the median preoptic nucleus (MnPO). Inhibitory responses (n=7) were not affected. Microinjection of ANG II into the region of the SFO produced either a facilitation (n=28) or no effect (n=6) on the excitability of phasically active PVN neurosecretory cells and the facilitatory effect of 9 out of 23 cells tested was prevented by pretreatment with Sar in the region of the MnPO. All the PVN cells which had excitatory responses to either electrical (n=7) or chemical (n=9) stimulation of the SFO that were blocked following the pretreatment could also be activated by intravenous administration of ANG II. Furthermore, this activation was blocked (n=10) or attenuated (n=6) by pretreatment with Sar in the region of the MnPO. These results show an involvement of both the MnPO and the SFO for the regulation of excitability of putative vasopressin (VP)-secreting PVN neurons, and suggest that MnPO neurons sensitive to ANG II may relay activation of SFO neurons by circulating ANG II to putative VP-secreting PVN neurons which result in enhanced excitability.     

A direct neural projection from the nucleus of the solitary tract to the subfornical organ in the rat

A direct neuronal projection from the nucleus of the solitary tract (NTS) to the subfornical organ (SFO) in rats was demonstrated using the Phaseolus vulgaris leucoagglutinin (PHA-L) anterograde tracing technique. Small deposits of PHA-L were iontophoretically placed within the caudal parts of the nucleus of the solitary tract. Brain sections were processed for visualization of PHA-L using the avidin-biotin immunoperoxidase technique. PHA-L immunoreactive beaded axons were observed within the subependymal and central parts of the SFO. The results demonstrate an anatomical substrate for direct feedback from cardiovascular regions of the NTS to the SFO. This pathway may be important for relaying blood pressure and associated body fluid volume changes to the SFO.     

A method for simultaneously recording neural activity and rotation in the rat

An apparatus is described which allows the simultaneous recording of neural activity and rotational behaviour in the rat. Rotational behaviour is measured, coded and stored on magnetic tape along with neural activity from chronically implanted electrodes, allowing off-line computer analysis.     

The A1 noradrenergic region enhances the responsivity of hypothalamic paraventricular neurohypophyseal neurons to inputs from the subfornical organ in the rat

Summary The action of the A1 noradrenergic neurons of the ventrolateral medulla on the responsiveness of neurohypophyseal neurons in the rat hypothalamic paraventricular nucleus (PVN) to inputs from the subfornical organ (SFO) was examined in antidromically identified PVN neurons that respond to electrical stimulation of both the SFO and A1 region. In both putative vasopressin (VP)- and oxytocin (OXY)-secreting PVN neurons that were classified according to their spontaneous firing patterns and their responsivity to baroreceptor activation, prior stimulation of the A1 region did not affect the short latency brief duration excitatory response induced by SFO stimulation. Simultaneous stimulation of the A1 region significantly enhanced the long latency prolonged excitatory response induced by SFO stimulation and the enhancement was blocked by microiontophoretically applied phentolamine, and -adrenoceptor antagonist, but not by timolol, a -adrenoceptor antagonist. Simultaneous stimulation of the A1 region also significantly enhanced the inhibitory response induced by SFO stimulation and the enhancement was blocked by microiontophoretically applied timolol, but not by phentolamine. These results suggest that the A1 region may act to enhance the partial excitatory (via an -adrenoceptor mechanism) and inhibitory SFO inputs (via a -adrenoceptor mechanism) to the PVN neurohypophyseal neurons as a modulatory action.