Effects And Mechanism Of Supraoptic Nucleus On Gastric Function In Rats

Hypothalamus is the center of neuroendocrine which combines the neuromodulation and humoral regulation to modulate the visceral function, such as body temperature, feeding, reproduction, humour osmotic pressure, endocrine and so on. It is composed with preoptic region, supraoptic region, tuberal region and bulla fornicis region from rostral to caudal end. Supraoptic nucleus(SON) and paraventricular nucleus(PVN) are the most prominent nuclei in anterior hypothalamus, their neuronal morphology and physiology are similar in Golgi-stained or immunohistochemistry studies, both of which are composed of vasopressinergic(VP) neurons and oxytocinergic (OT) neurons.It is well known that both of the SON and PVN neurons synthesize VP and OT, and then transport and store in neurohypophysis via supraopticohypophyseal tract or paraventriculohypophyseal tract. VP and OT will be released and combine with target organ, for example, when the osmotic pressure of plasma increase, lactate or deliver. The target organs of VP are located in distal convoluted tubule and collecting tube, which will affect reabsorption and plasma osmotic pressure, and the target organs of OT are located in uterus and mammary gland, which will affect delivery and lactation.However, more and more evidences confirm that there are functional connections between PVN and the stomach. Rogers et al found that stimulation of PVN reduced gastric acid secretion and inhibited gastric motility, and the latter standpoint was supported by Sakaguchi. Humphreys et al reported that injection of neuropeptide Y into the PVN inhibited gastric acid secretion in the rat.Gastric motility and food intake elevated after lesions of PVN in rats. The effects of PVN on stomach can be abolished by bilateral subdiaphragmatic vagotomy, so we can conclude that the PVN modulates the stomach via vagal parasympathetic nerves which come from the neurons located in dorsal motor nuclei of vagus ( DMV ) and partly in nucleus ambiguus(NA), and Sofromew et al confirmed there were direct projections from OT and VP neurons in PVN to DMV via immunohistochemical visualization of the horseradish peroxidase and neuroregulator. So far, whether there are functional connections between SON and stomach remains unknown. In other studies, gastric secretion increased after direct microinjecting methacholine chloride, a kind of incentive chemical stimulation, into the SON in dogs, and this phenomenon could be eliminated after bilateral vagotomy. Ueta et al. consistently observed that discharge of VP neurons were inhibited by gastric distension which were induced by balloon inflation of 4-8 ml in rat. Takayuki discovered the gastric distention of physiological levels stimulated the vascular release of 5-HT, and induced c-Fos expression (represent neuronal activity) in the NTS, area postrema (AP), PVN and SON, and could be blocked by truncal vagotomy, thence he presumed that there were fiber projections between NTS and SON.Our prior studies investigated the c-Fos expression in medulla oblongata and hypothalamus nuclei during different durations (30,60,120,180min) of restraint water-immersion stress(RWIS) in cold water(21±1℃) in the rat. The most intense c-Fos induction was always observed in the SON and the PVN in hypothalamus, and as well as DMV, NA and NTS in medulla oblongata. These data strongly suggest that SON, PVN, DMV, NA, NTS participate in the disorders of gastric motility and secretion induced by the RWIS, and many evidences indicate that there are functional connections between all of the nucleus mentioned above and stomach except for SON.On the whole, these reports and our prior work indicate that there are functional connections btween the SON and stomach. On earth, whether the connections exist and the pathway remains unknown. Now that, the present study will resolve it.Firstly, whether there are bidirectional functional connections between SON and stomach. The rats were anesthetized and then divided into four groups randomly. Group I, the neuronal discharge was recorded extracellularly in the NTS, DMV or SON before and after the cold physiologic saline (4℃) was perfused into the stomach from esophagus and effused from the duodenum for 5 minutes, the current was fluent so as to avoid gastric distension. Group II, the rats were stimulated by cold physiologic saline as Group I , 60 minutes later, the c-Fos expression of the NTS, DMV and SON was examined via immunohistochemistry technique, respectively. Group III was the control to Group II and was not perfused with cold physiologic saline (4℃). Group IV, gastric motility was recorded continuously by inserting a latex balloon(volume 0.2 ml, pressure 110 mmHg) into pylorus through the forestomach which connected with pressure transducer. In turn, connected to the BL-420F. Then the amplitude, duration, frequency, and motility index of gastric contraction waves within 10 minutes before and after microinjection of L-Glu(0.1mol/L,50 nL) into the SON were measured. Results: In Group I,the discharge frequency caused by the gastric lower temperature increased in all the three nuclei, but differed in degree. DMV and SON had significant difference (P <0.05) than before the perfusion of cold water (For example, SON, 6.85±1.71 spikes/s vs 4.5±0.68 spikes/s, n=7), but NTS had no significant difference. In Group II, c-Fos expression in NTS, DMV and SON was respectively greater than that of Group III (P<0.05)(e.g. in SON, the number of Fos-positive neurons is 188.23±16.45 vs 102.82±7.39, n=5). In Group IV, microinjection of L-Glu(5 nmol, n=7) into SON significantly inhibited gastric motility, motility index before injection was 1482.1±708.2 and 1185.5±639.2 after injection (P <0.05). These data strongly suggest that there are functional connections between supraoptic nucleus and stomach in rat, and imply that the SON may modulate gastric motility via activating DMV, which is similar with the PVN.Secondly, effects of electrical stimulation or L-Glu microinjected into SON on gastric motility. The results demonstrate that the gastric motility amplitude and index was significantly inhibited by 0.1, 0.01, or 0.001 mol/L L-Glu, the inhibitory rate was 21.3%,20.6% and 17.9%, there are dose dependent manner btween L-Glu and gastric motility; but not the respiratory, heart rate and blood pressure. The inhibitory effects could be abolished by diaphragmatic vagotomy, hexamethonium bromide (0.5 ml/100g,15 mg/Kg)or acetylcholine injection (Ach, 0.5 ml/100g,600μmol/Kg). These results suggested that SON modulate stomach via the efferent pathway of vagal nerves, and activating the cholinergic preganglionic neurons. The electrical stimulation,0.15 mV, of SON could inhibit the gastric motility and the inhibitory rate was 23.5%.Thirdly, effects of electrical stimulation or L-Glu microinjected into SON on gastric acid secretion.The physiologic saline (2.0 ml/min, pH7.4, 37℃) was perfused into the stomach from esophagus and effused from the duodenum, the current was fluent so as to avoid gastric distension. Take 20 min as a unit to collect the gastric acid secretion, and compared the after stimulation versus before. Results, gastric acid secretion increased by 0.1, 0.01 or 0.001 mol/L L-Glu microinjecting into the SON, and the increasing effect maximized at 3 hours and lasted for 4 hours later, as well, the effects could be abolished by diaphragmatic vagotomy. Electrical stimulation SON could enhance the gastric acid secretion and maximized at 1.5 hours. Lastly, the pathway and some more related studies. The discharges of diaphragmatic vagus nerve increased signifcantly under the condition that SON was excited by 0.1 mol/L L-Glu microinjecting, however, there were no Fos-like immunoreactive neurons in preganglionic neurons of sympathetic nerve innervating the stomach. Besides, we confirmed that the frenquency of gastric myoelectricity was more important than amplitude, and it could be increased when SON was excited by 0.1 mol/L L-Glu microinjecting.In conclusion, the present studies comfirmed the functional connections between SON and the stomach in rats, and we can found that:1. The increase of discharge and c-Fos expression under the stimulus (gastric temperature decreased instead of rectum ) indicated that the gastric wall thermoreceptors excited and, in turn, activated the DMV, NTS and SON. vice versa, the SON could modulate the stomach via vagus nerve.2. When the SON was excited by electrical stimulation or L-Glu microinjecting, the gastric function was affected, however, the gastric motility decreased and the gastric acid secretion increased, they were not synchronous.3. The sympathetic nerve did not involve in the functional modulation between SON and stomach.4. Gastric myoelectricity may be as useful as electrocatdiogram and electroencephalogram in the future.In a word, we have obtained evidence of functional connections between SON and stomach, thence, more data about mechanism of gastric modulation and function of hypothalamus SON will be derived from our experiment.