Stimulation Of Median Eminence Research Articles

Mechanisms of Neuropeptide Y, Peptide YY, and Pancreatic Polypeptide Inhibition of Identified Green Fluorescent Protein-Expressing GABA Neurons in the Hypothalamic Neuroendocrine Arcuate Nucleus

The fast inhibitory transmitter GABA is robustly expressed in the arcuate nucleus (ARC) and appears to play a major role in hypothalamic regulation of endocrine function and energy homeostasis. Previously, it has not been possible to record selectively from GABA cells, because they have no defining morphological or physiological characteristics. Using transgenic mice that selectively express GFP (green fluorescent protein) in GAD67 (glutamic acid decarboxylase 67)-synthesizing cells, we identified ARC GABA neurons (n > 300) and used whole-cell recording to study their physiological response to neuropeptide Y (NPY), the related peptide YY(3-36) (PYY(3-36)), and pancreatic polypeptide (PP), important modulators of ARC function. In contrast to other identified ARC cells in which NPY receptor agonists were reported to generate excitatory actions, we found that NPY consistently reduced the firing rate and hyperpolarized GABA neurons including neuroendocrine GABA neurons identified by antidromic median eminence stimulation. The inhibitory NPY actions were mediated by postsynaptic activation of G-protein-linked inwardly rectifying potassium (GIRK) and depression of voltage-gated calcium currents via Y1 and Y2 receptor subtypes. Additionally, NPY reduced spontaneous and evoked synaptic glutamate release onto GABA neurons by activation of Y1 and Y5 receptors. The peptide PYY(3-36), a peripheral endocrine signal that can act in the brain, also inhibited GABA neurons, including identified neuroendocrine cells, by activating GIRK conductances and depressing calcium currents. The endogenous Y4 agonist PP depressed the activity of GABA-expressing neurons mainly by presynaptic attenuation of glutamate release. Together, these results show that the family of neuropeptide Y modulators reduces the activity of inhibitory GABA neurons in the ARC by multiple presynaptic and postsynaptic mechanisms.

Postsynaptic Effects of Prolactin and Estrogen on Arcuate Neurons in Rat Hypothalamic Slices

The effects of 17 beta-estradiol (E2) and prolactin (PRL) on the excitability of neurons in the arcuate nucleus were examined using hypothalamic slices obtained from ovariectomized rats. Neither E2 nor PRL affected the excitability of neurons having antidromic responses due to median eminence stimulation. On the other hand, E2 and PRL altered the spontaneous activity of unidentified neurons, which were not activated by median eminence stimulation, by 6 and 11%, respectively. Both excitatory and inhibitory responses were recorded, which were not abolished in the low Ca2+/high Mg2+ medium. These results suggest that E2 and PRL can rapidly modulate the excitability of some arcuate, but presumably not tuberoinfundibular, neurons by acting directly on the postsynaptic membrane.

Bromocryptine Prevents the Decline in Tuberoinfundibular Neuronal Release of Dopamine after Removal of Chronic Estrogen Treatment

Prolonged exposure to estradiol 17-beta (E2) in rats has been shown to decrease dopamine (DA) synthesis in and release from tuberoinfundibular dopaminergic (TIDA) neurons in Fischer 344 rats. The objective of the present study was to determine whether inhibition of the E2-induced increase in anterior pituitary (AP) weight and prolactin (PRL) secretion by concomitant administration of the dopaminergic agonist, bromocryptine, could prevent the decrease in TIDA neuronal function produced by chronic E2 administration. TIDA neuronal function was evaluated by in vitro superfusion and electrical stimulation of median eminence (ME) tissue after allowing for accumulation of [3H]dopamine (DA). The effect of chronic E2 and/or bromocryptine treatment on catecholamine content in tuberohypophyseal neurons in the neurointermediate lobe was also measured to determine whether increased pituitary size possibly damaged the tuberohypophyseal neurons. Treatment with E2 for 30 days significantly increased AP weight, serum PRL concentration, and AP PRL and DNA content over values in non-E2-treated controls. When bromocryptine was injected daily during E2 treatment, bromocryptine completely inhibited the E2-induced increase in serum PRL and AP DNA content, and AP weight was only moderately increased. The evoked release of 3H at the end of the 30-day E2 treatment was reduced during electrical stimulation and there was no augmented release of 3H from the ME tissue after 10 microM nomifensine infusion in E2-treated rats and in rats given both bromocryptine and E2. However, neurointermediate lobe DA content was diminished only in E2-treated rats and not in animals given bromocryptine together with E2. When all treatments were discontinued for 30 days, animals previously given only E2 showed sustained increases in AP weight, serum PRL levels, and AP PRL and DNA content, but reduced stimulation-evoked release of 3H, absence of response to nomifensine, and reduced neurointermediate lobe DA and norepinephrine content when compared with values in non-E2-treated controls. After withdrawal of E2 treatment for 30 days, animals previously given bromocryptine and E2 together were not different from control animals in any of the parameters measured. These results suggest that the decline in TIDA neuronal release of DA induced by chronic E2 treatment was at least partly exerted via the marked hyperprolactinemia and/or by compression of the medial basal hypothalamus by the enlarged AP.

Hypothalamo-hypophysial blood flow: a novel control mechanism in pituitary function?

The flow rate of portal plasma was measured during hypothalamo-hypophysial portal sampling in the rat using a modified Worthington-Fink technique. In rats rendered hypothyroid with propylthiouracil, there was a significant 140% increase in portal plasma flow. Median eminence stimulation also increased portal plasma flow by 41%. Orchidectomy and adrenalectomy had no effects on plasma flow. Modification of flow rate in the hypothalamo-hypophysial vascular bed may represent a further mechanism involved in the control of pituitary function.

Antidromic activation of rat dorsomedial hypothalamic neurons from locus coeruleus and median eminence

In rats anesthetized with urethane, single unit activity was recorded in the hypothalamic dorsomedial nucleus (DMH) to obtain antidromic response to stimulation of locus coeruleus (LC) and median eminence (ME). Ninety-two cells were activated antidromically from LC and/or ME. Antidromic latencies to LC stimulation ranged from 7 to 39 msec and those to ME stimulation ranged from 5 to 20 msec. Approximately 13% of the neurons recorded revealed antidromic responses simultaneously from LC and ME, and they were found to bifurcate near the soma. The majority of DMH neurons projecting to LC alone were not spontaneously active, while those projecting to ME tended to discharge spontaneously.

Responses of arcuate neurons to some putative neurotransmitters in perfused rat hypothalamic slices: effects of in vivo and in vitro estrogen treatments.

The responses of neurons in the arcuate nucleus (ARC) to gamma-aminobutyric acid (GABA), glycine, serotonin (5-HT), norepinephrine (NE), and dopamine (DA) were examined in hypothalamic slices from ovariectomized rats either treated or untreated with estrogen. In estrogen-untreated preparations, spontaneous unit activity of the majority of neurons identified antidromically by the median eminence (ME) stimulation was depressed by GABA and glycine. When effective, 5-HT was predominantly excitatory and DA was predominantly inhibitory, while NE evoked both excitatory and inhibitory responses. The IS-SD block in evoked spikes was also occasionally observed following GABA and glycine perfusion. The responsiveness of unidentified neurons, which were not activated by the ME stimulation, to agents other than 5-HT did not differ from that of antidromically identified neurons. Responses of unidentified neurons to 5-HT were predominantly inhibitory. Subcutaneous injection of 20 micrograms of 17 beta-estradiol-benzoate 2 days prior to the experiments did not significantly affect the responsiveness of both identified and unidentified neurons to GABA, 5-HT, DA, and NE. Contrarily, preincubation of slices in 17 beta-estradiol-containing medium (10 nM) for 1 h significantly increased the responsiveness of unidentified, but not identified, neurons to the inhibitory action of NE. These results suggest that GABA, glycine, 5-HT, NE, and DA play some functional roles in controlling the excitability of tuberoinfundibular (TI) neurons in the ARC. It is also suggested that the negative feedback effect of estrogen is mediated, at least in part, by increased responsiveness of ARC neurons other than TI neurons to the inhibitory action of NE.

Combined electrophysiological, immunocytochemical and peptide release measurements in the hypothalamic slice

Sagittal hypothalamic slices were prepared from cycling female guinea pigs and incubated for 10–12 h during which time intracellular recordings were made from 274 arcuate and cell-poor zone (ARC-CPZ) neurons. Many of the cells exhibited spontaneous activity and a small percentage could be driven antidromically via median eminence stimulation and orthodromically via stria terminals stimulation. During the long-term recording, the effluent medium was collected in 30 of the experiments and radioimmunoassayed using a high-titer, conformational antisera for the peptide luteinizing hormone-releasing hormone (LH-RH). The findings revealed that LH-RH is released in detectable amounts from slices prepared from a single hypothalamus, that the amount of LH-RH released is dependent on the stage of the cycle, and that it can be stimulated by known secretagogues like prostaglandin E 2. Following the electrophysiological experiments, the slices were fixed and processed for immunocytochemistry using a specific antisera (EL-14) for LH-RH. LH-RH neurons were stained throughout the ARC-CPZ area with intense fiber staining in the median eminence. These findings demonstrate that the hypothalamic slice preparation can be utilized for combined electrophysiological, immunocytochemical and peptide release measurements.

Effects of neural stimuli on paraventricular nucleus neurones

The electrical activity of 122 neurones mostly located within the parvocellular components of the hypothalamic paraventricular nucleus (PVN) was recorded in urethane-anaesthetized male rats. Spontaneous activity and responses to single-shock stimulation of the following sites were recorded: median eminence (ME), neurohypophysis (PIT), preoptic area (POA), nucleus accumbens (ACB), subiculum (SUB), sciatic nerve (ScN), and photic (Ph) stimulation. Stimulation of ME antidromically invaded 14% of the cells recorded, the spontaneous firing rate of these neurones was significantly slower than that of the rest of the population. The responses of the neurones identified as projecting to ME, to the above stimuli, were generally found to be in the same direction as for the whole population but the proportions of the majority responses were greater. Identified cells mainly responded with orthodromic excitation following POA, SUB, ScN, and Ph stimulation whereas ACB stimulation produced a greater proportion of inhibitiory responses. High degrees of convergence were found following afferent stimuli. The possibility that some of the neurones projecting to ME may be candidates for the corticotropin-releasing factor (CRF-) cells, is discussed.

Effects of hyperprolactinaemia and testosterone on the release of LH-releasing hormone and the gonadotrophins in intact and castrated rats.

We have investigated the effect of hyperprolactinaemia on the secretion of LH-releasing hormone (LHRH), LH and FSH in male rats of the PVG strain which were left intact, castrated or castrated and then implanted with either a 10 or 30 mm silicone elastomer capsule containing testosterone (T10 and T30 respectively). Hyperprolactinaemia was produced by pituitary grafts under the kidney capsule. Pituitary stalk blood, for LHRH estimation, and peripheral blood, for LH, FSH and prolactin, were collected under alphaxalone anaesthesia. Pituitary stalk blood was collected during three consecutive periods of 30 min each before, during and after the application of an electrical stimulus to the median eminence (ME). Hyperprolactinaemia significantly reduced the plasma concentrations of FSH in intact rats and the post-castration increase in the plasma concentrations of both LH and FSH. Neither hyperprolactinaemia nor castration had any significant effect on the spontaneous output of LHRH, but castration alone or castration plus implantation of a T30 capsule did significantly reduce the increment in LHRH output produced by ME stimulation, an effect not seen in rats bearing pituitary grafts. The T30, but not the T10 capsules suppressed the post-castration increase in the gonadotrophins, and the inhibitory effect of testosterone was not significantly affected by hyperprolactinaemia. An incidental but important finding was that the presence of pituitary grafts under the kidney capsule reduced the anaesthetic dose of alphaxalone by 63%.(ABSTRACT TRUNCATED AT 250 WORDS)

A hypothalamic-pituitary system that stimulates the release of plasminogen activator in the rat

We have studied the possible role of the hypothalamic-pituitary system in the control of the release of plasminogen activator (PA) into peripheral blood of male rats. Plasminogen activator was measured by euglobulin lysis time. Desamino- d-arginine 8 vasopressin (dDAVP) and adrenaline injected i.v. induced an increase in plasma PA as did electrical stimulation of the median eminence (ME), but dDAVP had no effect on plasma PA in hypophysectomized rats. The PA response to ME stimulation was similar in Brattleboro rats (deficient in vasopressin) and adrenalectomized Wistar rats compared with intact Wistar rats, but was abolished by section of the pituitary stalk and was negligible in hypophysectomized rats. The 41-residue corticotropin releasing factor (CRF) had no effect on PA release. Saline extracts of anterior pituitary gland from both normal Wistar and Brattleboro rats produced a dose-dependent increase in plasma PA when injected into normal Wistar rats. The activity of pituitary tissue was abolished by boiling, but not by di-isopropyl fluorophosphate which inactivates PA itself. Thus the anterior pituitary gland of the rat contains a heat-labile factor which stimulates the release of PA from peripheral stores into the circulation. This pituitary factor is released by a hypothalamic factor that is neither vasopressin nor CRF.

In vitro neurophysiology of identified rat hypothalamic 'neuroendocrine' neurons.

Stable synaptic and antidromic electrophysiological recordings were obtained for 10-15 h from identified neurohypophyseal and tuberoinfundibular neurons using an acute explant of rat hypothalamus maintained in vitro through perfusion of the carotid artery with oxygenated artificial media. Synaptic potentials from preoptic and median eminence stimulation were blocked during perfusion with 12 m M Mg++-containing solutions. Antidromic and spontaneous action potentials recorded in neurohypophyseal neurons were followed by a transient after-hyperpolarization of 3-10 mV, due primarily to a brief increase in potassium conductance. Evidence favoring the existence of a synaptic recurrent inhibitory pathway was present in records obtained from tuberoinfundibular neurons, but not from neurohypophyseal cells.

Endogenous GABA receptor ligands in hypophysial portal blood.

The concentrations of endogenous gamma-aminobutyric acid (GABA) receptor ligands were measured in hypophysial portal plasma by a sensitive and specific radioreceptor assay. Portal plasma contained higher concentrations of GABA receptor ligands than peripheral controls, although this increase was not due to authentic GABA but an unknown low-affinity ligand. Evidence is presented to support the hypothesis that the dipeptide homocarnosine may represent at least a part of this unknown. Electrical stimulation of median eminence evoked a massive (7.8-fold) increase in the rate of release of GABA (but not the low-affinity component) into portal blood. These results suggest that GABA and possibly other low-affinity ligands such as homocarnosine could serve as factors involved in the central control of anterior pituitary function.

Initial report on combined in vivo single cell recording and intracellular staining

This paper reports preliminary data obtained from the combination of extracellular single unit recording, microiontophoretic testing and intracellular staining of single neurons in the diencephalon of the anesthetized rat. Sixty neurons were recorded extracellularly and iontophoretically tested with glass multibarrelled micropipettes. Thirty-four of these neurons were identified by antidromic invasion from median eminence stimulation. Seventeen of these antidromically identified neurons were subsequently impaled with the micropipette and intracellularly stained with the fluorescent dye lucifer yellow-CH. The average diameter of the antidromically identified neuronal cell bodies was 7.4 microns. The iontophoretic response profiles of these stained neurons were similar to the profiles of non-stained antidromically identified neurons. Four of the remaining 26 neurons were synaptically activated from median eminence stimulation and were successfully marked with lucifer yellow. Average soma diameters of these neurons was 12.8 microns. Twenty-two neurons were not antidromically or orthodromically identified from median eminence stimulation. Three of these neurons were intracellularly stained with lucifer yellow and their soma diameters averaged 6.9 microns. Approximately 50% of all staining attempts, subsequent to extracellular recording and iontophoretic testing, were successful. The combination of these techniques is therefore a feasible approach to the in vivo study of the physiologic, pharmacologic and morphologic properties of single neurons.

Connections of the hypothalamic paraventricular necleus with the neurohypophysis, median eminence, amygdala, lateral septum and midbrain periaqueductal gray: An electrophysiological study in the rat

Extracellular recordings were obtained from 555 paraventricular (PVN) nucleus neurons in pentobarbital-anesthetized male rats. Cells were examined for their spontaneous activity patterns and response to single 1-Hz electrical stimulation of the neurohypophysis, median eminence, amygdala, lateral septum (LS) and midbrain periaqueductal gray (PAG). Neurohypophyseal stimulation evoked antidromic activation from 109 neurons. Among spontaneouly active neurohypophyseal neurons, evidence of a recurrent inhibitory pathway usually required pituitary stimulus intensities twice threshold for antidromic activation. Orthodromic excitatory or inhibitory responses followed amygdala and LS stimulation, but not PAG stimulation. The amygdala influence was predominantly inhibitory to ‘phasic’ (putative vasopressin-secreting) PVN neurohypophyseal neurons. Neurohypophyseal stimulation evoked orthodromic responses from 124 PVN cells; some of these neurons were also responsive to stimulation in other sites. Median eminence stimulation evoked antidromic responses from 37 PVN neurons; some of these cells also displayed phasic activity but not evidence for recurrent inhibition. Twelve cells in this group were also activated antidromically from both the median eminence and the neurohypophysis; collision tests suggest that the median eminence innervation may be an axon collateral of a neurohypophyseal pathway. Amygdala stimulation was inhibitory to some cells in this category. Amygdala, LS and PAG stimulation evoked antidromic activation from a small number of PVN cells, but none of these cells appeared to innervate more than one area, including the neurohypophysis, and none displayed phasic activity. Orthodromic responses were recorded among other PVN neurons after stimulation in these sites; however, PAG stimulation was the least effective stimulation area. These observations provide additional electrophysiological data that confirm efferent PVN connections to all areas tested, afferent connections from amygdala and LS but not PAG, and the possibility for coordinated activity among PVN neurons through local recurrent or common afferent connections.

Stimulation-dependent changes in synaptic effects on unit activity of medial preoptic nucleus neurones in rats.

Stimulation-dependent changes in synaptic effects were observed in medial preoptic nucleus neurones during stimulation of the amygdala or pyriform cortex in anaesthetized female rats. The changes occurred after 35--240 triple pulse stimuli repeated at 0.89 Hz. Median eminence stimulation did not produce any synaptic change. These data show the existence of synaptic plasticity in the neural pathway from the amygdala and pyriform cortex to the medial preoptic nucleus.

Amygdala neurones: converging synaptic inputs produced by median eminence and medial preoptic area stimulations in rats.

1. Amygdaloid afferent inputs from the median eminence and the medial preoptic area were studied electrophysiologically in urethane-anaesthetized female rats.2. Stimulation of the surface of the median eminence produced orthodromic responses in about 80% of the forty-seven amygdala units tested and about 65% of the responsive cells showed an excitation.3. Stimulation of the ipsilateral medial preoptic area orthodromically excited seventeen and inhibited twenty of the forty-nine units tested.4. Stimulation of the contralateral medial preoptic area evoked orthodromic excitation in nine and inhibition in three of the twenty-two units tested. These stimuli were ineffective for producing antidromically conducted impulses in the tested units.5. Sixteen of the thirty amygdala units tested for responses to both median eminence and ipsilateral medial preoptic area stimulation responded orthodromically with either excitation or inhibition. The latency of the response to median eminence stimulation was approximately equal to that of the response to ipsilateral preoptic area stimulation in four of these sixteen units.6. A characteristic bursting discharge was observed in eleven amygdala units during and after ipsilateral preoptic area stimulation. A transitory inhibition was evoked simultaneously with the bursting discharge in some units. Seven of the eleven units were also tested for median eminence stimulation, and a transitory excitation was observed in each of these units.7. These results suggest the existence of converging synaptic inputs both from tuberoinfundibular neurones and the ipsilateral medial preoptic area to certain amygdala neurones. They also demonstrate the existence of a specific neural pathway mediating a characteristic self-sustained bursting discharge in some amygdala neurones after such stimulation.

Influence of midbrain stimulation on the excitability of neurons in the medial hypothalamus of the rat

Extracellular action potentials were recorded from 1098 neurons in the medial hypothalamus of pentobarbital anesthetized male rats. Their excitability was analyzed after single 1 Hz stimulation of the midbrain periaqueductal gray (PAG) or adjacent reticular formation. Cells were also examined for their response to median eminence (ME), amygdala, lateral septum (LS) or anterior hypothalamic/preoptic area (AHA-POA) stimulation. Antidromic invasion from midbrain stimulation was recorded from 110 neurons. Eight of these neurons showed features of axon branching and displayed antidromic invasion from both midbrain and amygdala (2 cells) or AHA/POA (6 cells). Many neurons with midbrain projections displayed orthodromic responses to stimulation in the amygdala, but few responded to AHA/POA or LS stimulation. Midbrain stimulation evoked orthodromic responses from 99 medial hypothalamic neurons. Many of these cells also displayed orthodromic responses to amygdala or AHA/POA stimulation, whereas a small number were activated anti-dromatically by stimulation in these sites. None of 42 neurons activated antidromically from median eminence stimulation were responsive to midbrain stimulation. These results provide electrophysiological evidence of reciprocal connections between medial hypothalamic and medial midbrain areas, and indicate that medial hypothalamic neurons with midbrain connections are subject to influences from other extrahypothalamic areas.

Immunoreactive luteinizing hormone releasing factor (LRF) in pituitary stalk plasma from female rats: effects of stimulating diencephalon, hippocampus and amygdala.

1. Previous studies on the effect of preoptic and median eminence stimulation on the immunoreactive LRF content of pituitary stalk blood from pro-oestrous rats have been extended. Stimulation of the suprachiasmatic nuclei and anterior hypothalamic area produced increments in LRF which were 66 and 18% respectively, of that produced by preoptic stimulation, and 38 and 9%, respectively, of that produced by stimulation of the median emience. Stimulation of the amygdala and hippocampus had no effect. 2. The LRF response was not affected significantly when preoptic stimulation was accompanied by stimulation of the hippocampus. 3. In animals subjected to section of the dorsal afferents of the diencephalon, the LRF response to preoptic stimulation was similar to that in intact rats. However, the facilitatory effect of oestrogen on the LRF response to preoptic stimulation was significantly reduced in the roof sectioned compared with intact animals. The post-operative resumption of oestrous cycles was delayed but not abolished by dorsal deafferentation.

Tuberoinfundibular neurons in the basomedial hypothalamus of the rat: Electrophysiological evidence for axon collaterals to hypothalamic and extrahypothalamic areas

In pentobarbital or urethane anesthetized rats, the activity of 889 mediobasal hypothalamic neurons was studied for evidence of a response to median eminence stimulation. Evidence of antidromic invasion, which indicated a projection to the median eminence, identified 134 cells (15%) as 'tuberoinfundibular' neurons. Antidromic spike latencies ranged from 0.5 to 14.0 msec (4.3 +/- 2.9 S.D.); conduction velocities were under 1.0 m/sec and were generally slower for tuberoinfundibular neurons located closest to the ventral surface of the hypothalamus. Certain tuberoinfundibular neurons followed paired median eminence shocks at frequencies up to 500 Hz; an increase in both the threshold and the latency for the second antidromic spike was observed with interstimulus intervals under 4 msec. Only 38% of tuberoinfundibular neurons were spontaneously active; 24 of 29 spontaneously active neurons displayed evidence of recurrent inhibition with durations up to 150 msec and at latencies which approximated that of the antidromic spike but which did not depend upon antidromic invasion. Similar responses were observed from 33 spontaneously active non-tuberoinfundibular neurons. Evidence of orthodromic excitation in response to median eminence shocks was observed from 22 other medial hypothalamic neurons. Latencies for excitation ranged from 1.5 to 9.0 msec (mean 4.5 +/- 2.1 S.D.). Simultaneous antidromic invasion from other hypothalamic and extrahypothalamic sites was observed from 8 tuberoinfundibular neurons. These sites included the anterior hypothalamic area (2 cells), the preoptic area (3 cells) and the thalamic nucleus medialis dorsalis (3 cells). These results indicate the presence of axon collaterals within the tuberoinfundibular system; some appear to terminate locally within the hypothalamus, while others extend rostrally and dorsally into extrahypothalamic areas. These connections may provide pathways for extrahypothalamic distribution of peptides which regulate adenohypophyseal secretion, and suggest that these peptides may subserve alternate regulatory roles within the central nervous system.

Effects of Limbic stimulation and the microinontophoresis of aminergic drugs on the single unit activity of hypothalamic neurosecretory cells (author's transl).

Attempts were made to investigate the chemical nature of hypothalamic afferents from the extrahypothalamic structures which participate in the control of hypothalamic neurosecretory cells. Experimental procedures were discribed in the preceding report, except that of the electrical stimulation and the microinontophoresis. Some part of data obtained in this study were referred in the preceding report for improvement of statistic reliability. For electrical stimulation, side-by-side bipolar electrodes were placed in the medial preoptic area (MPO), medial amygdala (mAMYG) and dorsal hippocampus (dHPC) from the parietal surface of the brain and fastened to the cranium by dental resin. Microintophoresis of norepinephrine (NE), dopamine (DA) and acetylcholine (Ach) was carried out by using five-barrelled glass pipette on the antidromically activated units in the medical basal hypothalamus by the median eminence stimulation. Extracellular potentials were recorded by the central barrel of each electrode. One of the outer barrels was filled with physiological saline and used for control purpose. (1) Eelctrical stimulation of the MPO with train pulses of 0.2 msec duration and 300 muA intensity in 100 HZ for 5 sec, induced facilitation and inhibition in 26.4% and 9.4% of 106 tests, respectively. By the mAMYG stimulation with 500 muA, facilitation was seen in 18.8% and inhibition in 10.4%, of 96 tests. The HPC stimulation induced facilitation in 10.0% and inhibition in 23.3% of 90 tests. (2) Facilitatory effect of the MPO was most frequently seen in the ovariectomized and estrogen primed rats (58.8% of the tests). In contrast, the inhibitory effect of the dHPC was striking in the diestrus-I (40.0%). (3) Microiontophoresis of NE induced facilitation in 60.0% and inhibition in 10.6% of 66 identified cells. DA induced facilitation in 45%, and inhibition in 11.7% of 60 tests. Ach induced facilitation in 41.4%, and inhibition in 13.8% of 58 tests. (4) By NE infusion, facilitation was most commonly seen in the proestrus (70.6%). Inhibition was most readily elicited by Ach infusion on the diestrus-I (30%). Facilitative effect of DA was eminent in the diestrus-I (81.8%). (5) Successive administration of NE and DA on identical units revealed that in 56.0%, the units were responsive to only one agent and the other was effectless. 25.5% of the units responded to both of NE and DA. (6) The conceidence of effects, either facilitation or inhibition, between that of electrical stimulation of the microinontophoresis, was calculated on 46 units. The facilitatory effect of the MPO was most commonly mimicked by NE infusion (81.8%), and the inhibitory effect of the dHPC by Ach infusion (46.7%). NE also mimicked facilitatory effect of mAMYG stimulation (73.3%). (7) Intraventricular infusion of 20 mug of NE induced significant increase of serum LH (p less than 0.05 to saline) when measured by radioimmunoassay, whereas DA resulted in a decrease of LH (p less than 0.01).