Estrogen increases galanin immunoreactivity in hyperplastic prolactin-secreting cells in Fisher 344 rats.

Galanin is a widely distributed regulatory peptide which modulates the pituitary secretion of PRL and GH. Estrogen administration strongly stimulates galanin gene expression in the rat anterior pituitary. In adult female Fischer 344 rats, estrogen also induces hyperplasia of lactotropes. We used immunocytochemical analysis to assess the effects of estrogen on galanin-like immunoreactivity (Gal-IR) in the rat pituitary and hypothalamus during sc diethylstilbestrol (DES) implantation and after its removal at 30 days. In the anterior pituitary, DES implantation increased the portion of Gal-IR-containing cells from less than 2% in the control rats to 18.3% after 3 days of DES and 36% after 30 days. These changes paralleled the lactotrope hyperplasia exhibited in response to DES exposure. Ten and 30 days after removal of the DES capsules, the percentage of Gal-IR-containing cells in the anterior pituitary decreased to 6.3% and 1.5%, respectively. Colocalization studies revealed that Gal-IR-containing cells were predominantly lactotropes. Immunoelectron microscopy demonstrated that Gal-IR was concentrated in the Golgi region of these hyperplastic lactotropes and suggests that little of the synthesized galanin is secreted. The distribution of Gal-IR in the hypothalamus, median eminence, and neurohypophysis was unaffected by DES treatment. These data demonstrate that galanin is synthesized by hyperplastic pituitary lactotropes of Fischer 344 rats and that peptide accumulation is dependent on the presence of circulating estrogens. In contrast, neuronal galanin synthesis in the hypothalamus does not appear to be regulated by estrogen.

Immunohistochemical localization of corticotropin-releasing factor- and arginine vasotocin-like immunoreactivities in the brain and pituitary of the American bullfrog (Rana catesbeiana) during development and metamorphosis.

Immunoperoxidase staining for corticotropin-releasing factor (CRF) in the median eminence was sparce or absent from premetamorphic tadpoles, but increased dramatically by late prometamorphosis. Quantitative photometry revealed that CRF-like immunostaining material in the median eminence was most dense in metamorphic climax tadpoles. Arginine vasotocin (AVT)-like immunostaining material was visualized in perikarya of the magnocellular nucleus, with extensive fiber staining seen in the medial basal and infundibular hypothalamus as well as in the median eminence and pars nervosa of the pituitary. AVT-like immunoreactive perikarya were virtually absent in premetamorphic tadpoles, but their number increased greatly by Taylor-Kollros stage XII and continued to increase after this stage. Quantitative photometry revealed that AVT-like immunoreactivity in the pars nervosa increased greatly at Taylor-Kollros stage XII and remained intense after this stage. AVT-like immunoreactivity did not appear in the median eminence until Taylor-Kollros stage XVI. Localization of AVT-like immunoperoxidase staining around portal vessels in the median eminence suggests an anatomical mechanism for delivery of AVT to anterior pituitary corticotropes. These results indicate that both CRF and vasotocinergic neuronal systems develop just before the activation of interrenal steroidogenesis which occurs during the later stages of metamorphosis in this species.

Immunohistochemical identification of galanin and growth hormone-releasing factor-containing neurons projecting to the median eminence of the rat.

Through the combined demonstration of the retrograde transport of True blue and the immunohistochemical staining of galanin (GAL), the GAL neurons that project to the median eminence were identified. Moreover, the distribution of GAL and growth hormone-releasing factor (GRF) was analyzed in the arcuate nucleus (ARC) with an elution-restaining procedure. Following the injection of True blue into the median eminence, GAL-positive cells labeled with True blue were found mainly in the ARC. In addition, a few GAL neurons in the periventricular, the paraventricular and the supraoptic nuclei were labeled with True blue. The elution-restaining results revealed that many retrogradely labeled GAL neurons in the ARC contained GRF. These findings suggest that a subpopulation of GAL neurons in the ARC sending axons to the median eminence produces, stores and releases a GRF-like peptide.

Comparative immunohistochemical study of the distribution of neuropeptide Y, growth hormone-releasing factor and the carboxyterminus of precursor protein GHRF in the human hypothalamic infundibular area.

It is now well documented that various polyclonal antisera to the human growth hormone-releasing factor (hGHRF, somatocrinin) visualize in the brain by immunohistochemistry the classical hypothalamic hypophysiotropic neurons and also antigens present in otherwise characterized peptidergic neuronal systems. The nature of these antigens is still an open question. One of these hGHRF antisera, raised against an immunogen of hGHRF1-44NH2, labels in the arcuate nucleus of the human mediobasal hypothalamus the neuropeptide Y (NPY) containing neurons which for the most part constitute a tuberoextrainfundibular system. The identity of the hGHRF-like substance present in these neurons with true somatocrinin has been assessed by performing a comparative immunohistochemical study including sequential double and triple labeling using the antiserum to hGHRF1-44NH2 in conjunction with antisera to the carboxyterminus of preprosomatocrinin (CTPG) and to NPY. This made it feasible to dissociate the hGHRF1-44NH2-immunoreactive neurons into two major subpopulations costaining either for CTPG of NPY, and a minor neuronal group displaying simultaneously the three labelings. A subset of arcuate neurons also showed NPY staining only. These results suggest that (1) the hGHRF-like antigen present in the majority of the NPY neurons is not true somatocrinin, or alternatively that preprosomatocrinin undergoes a unique maturational processing in these neurons, and (2) a subset of tuberoinfundibular somatocrininergic neurons produces and releases NPY which may be involved in the multifactorial control of the pituitary function.

Topographical relationships between catecholamine- and neuropeptide-containing fibers in the median eminence of the newt, Triturus alpestris. An ultrastructural immunocytochemical study.

Dopaminergic and peptidergic nerve fibers were simultaneously demonstrated with a double-labeling technique at the ultrastructural level. The first antibody, raised against tyrosine hydroxylase, was applied during the preembedding phase and visualized with the peroxidase method. The second antibody, raised against one of the peptides met-enkephalin, somatostatin or gonadotropin-releasing hormone (GnRH), was applied to the ultrathin sections and visualized with gold-labeled goat anti-rabbit IgG. The fibers of both categories were present in the zona externa of the median eminence, frequently contacting the basal lamina of the portal vessels. In addition, topographical relationships between different types of nerve fibers were observed in the perivascular areas, although there were no morphological signs of synaptic specializations. Using serial sections, it could be established that one GnRH-fiber contacted both a dopaminergic fiber and a fiber immunoreactive for met-enkephalin. The observations support earlier physiological data concerning the regulation of the hypothalamo-hypophyseal axis, with special emphasis on the release of neurohormones in the median eminence of the newt.

Sexual differentiation of growth hormone feedback effects on hypothalamic growth hormone-releasing hormone and somatostatin.

To investigate possible sex differences in the feedback regulation of growth hormone (GH) secretion, concentrations of immunoreactive GH-releasing hormone (GRF) and somatostatin (SS) were measured in the median eminence (ME) and the hypothalamus of male and female rats bearing the MtTW15 tumor, which secretes high amounts of GH and prolactin (PRL). Four weeks after tumor implantation in male rats, the GRF concentration in the whole hypothalamus, including the ME, was decreased by 37% (0.29 +/- 0.02 vs. 0.46 +/- 0.02 ng/mg protein in intact male controls; p less than 0.001) and the concentration of SS was increased by 40% (11.5 +/- 0.7 vs. 8.1 +/- 0.3 ng/mg protein in male controls; p less than 0.01). In female rats, the presence of tumor for 4 weeks caused a smaller (18%) reduction in GRF concentrations (0.27 +/- 0.02 vs. 0.33 +/- 0.03 ng/mg protein in intact female controls; p less than 0.05) and no significant change in SS concentrations (10.2 +/- 0.08 vs. 9.7 +/- 0.8 ng/mg protein in female controls). Tumor-related changes in GRF and SS concentrations were also more pronounced in male rats than in females, when determined separately in the microdissected ME and in the remaining hypothalamus. These differences occurred despite similar increases in serum GH, PRL and insulin-like growth factor I concentrations in male and female tumor-bearing rats. To assess which hormone (GH or PRL) was responsible for these changes, intact male rats were treated for 10 days with 2 daily s.c. injections of rat GH (rGH; 100 and 250 micrograms/day), rat PRL (100 and 250 micrograms/day) or vehicle.(ABSTRACT TRUNCATED AT 250 WORDS)

Galanin-like immunoreactivity is influenced by estrogen in peripubertal and adult rats.

Galanin gene expression in the anterior pituitary is potently stimulated by estrogen in adult rats. To evaluate the influence of estrogen on galanin during the peripubertal period 30- to 32-day-old female rats were treated with pregnant mare serum gonadotropin (PMSG, 10 IU s.c., 10.00 h). Galanin-like immunoreactivity (galanin-LI) in hypothalamic and pituitary tissues was evaluated 1, 2 or 3 days after PMSG treatment between 17.00 and 19.00 h. The PMSG treatment stimulated 17 beta-estradiol secretion, which induced a midafternoon LH surge 2 days after the PMSG treatment. Concentrations of galanin-LI at the time of this LH surge were elevated 82% in the anterior pituitary and 58% in the hypothalamus (without the median eminence) when compared to saline-treated female rats. On the 3rd day after the PMSG injection, galanin-LI was increased 236% in the anterior pituitary, 88% in the neurointermediate lobe and 39% in the median eminence compared to saline-treated female rats. These changes in galanin-LI were not observed in similarly aged male rats or ovariectomized rats treated with PMSG. In adult male rats, daily injections with 17 beta-estradiol valerate (10 micrograms/daily s.c.) for 1 week increased galanin-LI in the median eminence and neurointermediate lobe to an extent similar to that seen in juvenile female rats following PMSG treatment. In contrast, the high serum levels of 17 beta-estradiol achieved after 17 beta-estradiol valerate treatment increased galanin-LI in the anterior pituitary 65-fold. These studies indicate that galanin-LI is influenced by estrogen in peripubertal and adult rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Stress-induced suppression of the prolactin afternoon surge in ovariectomized, estrogen-treated rats and the nocturnal surge in pseudopregnant rats are accompanied by an increase in median eminence dihydroxyphenylacetic acid concentrations.

Experiments were performed to determine whether the suppression of prolactin (PRL) surges during restraint was accompanied by changes in the activity of tuberoinfundibular dopamine (TIDA) neurons in the median eminence. Animals were either ovariectomized and estrogen-treated (OVX-PEP) or cervically stimulated to induce pseudopregnancy (PSP). Restraint stress was administered by tying the hind legs together with plastic-coated bell wire. Animals were decapitated following 15 or 30 min of restraint stress or immediately after removal from the animal room (control) when PRL levels were basal (10.00 h), at the peak of the afternoon PRL surge in OVX-PEP animals (17.00 h) or the nocturnal PRL surge in PSP animals (05.00 h). Median eminence dopamine (DA) and dihydroxyphenylacetic acid (DOPAC) were significantly decreased in control rats at 17.00 h when compared to control rats at 10.00 h (103.1 +/- 3.7 vs. 85.8 +/- 3.3 and 11.4 vs. 7.1 +/- 0.4 pg/micrograms protein, respectively) and plasma PRL was markedly elevated. Restraint stress at 10.00 h resulted in a significant increase in serum PRL, but this increase was not accompanied by a change in DA or DOPAC when compared to control animals (103.1 +/- 3.7 vs. 107.9 +/- 4.8 and 11.4 +/- 0.4 vs. 10.4 +/- 0.6 pg/micrograms protein, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

[Ultrastructural features of somatostatin-containing neurons of the hypothalamus of rats with protein insufficiency]. / Ul'trastrukturnye kharakteristiki somatostatinsoderzhashchikh neironov gipotalamusa krys pri belkovoi nedostatochnosti.

The electron microscopic investigation was performed to analyze somatostatin-contained nerve terminals in the median eminence of 21 days old malnourished rats' hypothalamus. In nerve terminals of malnourished animals in compared with controls ones there was found the increased density of granular vesicles (11.62 +/- 0.40 and 8.56 +/- 0.39 in 1 micron2, respectively) and decreased density of electron lucent vesicles with 120-160 nm diameter (1.66 +/- 0.18 and 3.43 +/- 0.26 in 1 micro2, respectively). The revealed increase in density of granular vesicles in axon terminals with positive immunohistochemical reaction to somatostatin in malnourished rats was explained by slow somatostatin release.

Angiotensin II antiserum decreases luteinizing hormone-releasing hormone in the median eminence and preoptic aea of the rat

Ovariectomized female rats were sacrificed 3 h after intracerebroventricular microijnjection of normal rabbit serum (NRS), specific antiserum against angiotensin II (AB-AII) or atrial natriuretic factor (AB-ANF). AB-AII decreased plasma LH by 50% and LH-RH content by 70% in the median eminence and medial preoptic area, respectively, but did not change plasma FSH when compared to animals which receivede NRS. There was no difference in these parameters when the animals were injected with AB-ANF or NRS.These results indicate that endogenous AII plays a physiological role in LH release acting directly or indirectly through LH-RH neurons of the median eminence and medial peroptic area

Exogenous triiodothyronine lowers thyrotropin-releasing hormone concentrations in the specific hypothalamic nucleus (paraventricular) involved in thyrotropin regulation and also in posterior nucleus.

Recent evidence indicates that thyroid hormones can regulate thyrotropin secretion in vivo in part by inhibiting thyrotropin-releasing hormone (TRH) secretion itself. Therefore, to explore whether triiodothyronine (T3) interacts with the specific hypothalamic area involved in thyrotropin (TSH) secretory regulation, the paraventricular nucleus (PVN), Palkovitz micropunches from eight nuclear regions were obtained from 1,000-microns frozen coronal brain slices for immunoassay determinations of TRH. Rats were treated either with parenteral L-T3 for 6 days to induce experimental thyrotoxicosis, or 0.15 M saline. The induction of thyrotoxicosis was confirmed by demonstrating that mean plasma TSH concentrations fell from 108 to less than 10 microU/ml (p less than 0.01). TRH concentrations in the PVN were reduced concomitantly after L-T3 from 1.9 to 1.1 ng/mg protein (p less than 0.05). No reductions in TRH concentrations during T3 treatment occurred in other nuclear groupings except in the posterior hypothalamic nucleus. Total TRH content in the median eminence declined also in T3-treated animals from 1.77 to 1.29 ng, representing a 32% reduction (p less than 0.01). No significant change was seen in the median eminence content of the TRH structurally related dipeptide, cyclo(His-Pro). The data herein indicate that experimental thyrotoxicosis in the rat is associated with a selective reduction in TRH concentrations in the PVN, documenting T3 effects upon hypothalamic TRH metabolism per se.

Alterations in concentrations of dihydroxyphenylacetic acid in the median eminence of rats euthanatized with pentobarbital.

Concentrations of dopamine (DA) and one of its major metabolites, dihydroxyphenylacetic acid (DOPAC), were determined in selected brain regions of rats that were euthanatized either by decapitation or by intravenous injections of pentobarbital or Fatal Plus, a commercial preparation that contains pentobarbital. When compared with values in decapitated brains, pentobarbital increased the concentration of DOPAC in the median eminence, which contains terminals of tuberoinfundibular dopaminergic (TIDA) neurons. Fifteen minutes of restraint reduced the concentration of DOPAC in the median eminence of rats killed by decapitation or by injections of pentobarbital, indicating that pentobarbital does not mask restraint-induced decrease in TIDA neuronal activity. In contrast, none of the manipulations altered DA or DOPAC concentrations in the striatum, which contains terminals of nigrostriatal dopaminergic neurons. Thus, changes in the concentrations of DOPAC in the median eminence (an index of TIDA neuronal activity) induced by stress can be detected in rats euthanatized by either decapitation or an injection of pentobarbital.

Tyrosine hydroxylase in the stalk-median eminence and posterior pituitary is inactivated only during the plateau phase of the preovulatory prolactin surge.

This study examined changes in the activity of tyrosine hydroxylase (TH) in the stalk-median eminence (SME) and posterior pituitary (PP) during the preovulatory PRL surge. Immature female rats were injected with PMSG on day 28. Blood PRL levels were low on the morning of day 30, rose to a peak from 1400-1600 h, remained at a lower plateau from 1800-2400 h, and declined to basal levels on the morning of day 31. SME, PP, and striatum were removed from PMSG-treated rats at selected times during the periovulatory period and from age-matched control rats. TH activity was determined in tissue homogenates by a coupled hydroxylation-decarboxylation assay. Apparent Km and maximum velocity values with respect to 6-methyl tetrahydropterine were estimated from substrate saturation curves. The kinetic parameters for TH in either the SME or the PP of control rats were similar at 1100 and 1800 h on day 30. However, the apparent Km in both tissues was significantly lower than that in the striatum. The affinity of TH in the SME and PP was unchanged before and during the peak phase of the PRL surge, reduced significantly during the late plateau, and returned to presurge levels in the morning of day 31. TH activity in the striatum was similar at all times examined. To determine the state of activation of the enzyme, tissue homogenates were preincubated with cAMP, ATP, and magnesium. TH activity in the SME during the peak phase was unchanged by cAMP, and that in the PP was modestly increased. The relatively inactive enzyme in both tissues during the plateau phase was markedly activated by a cAMP-dependent mechanism. The low affinity of striatal TH was greatly increased by cAMP at both times. These data suggest that TH in the SME and PP exists in an activated state most of the time and is transiently inactivated during the plateau phase of the PRL surge. In contrast, TH in the striatum is relatively inactive in the basal state and is not affected by hormonal changes induced by PMSG. We conclude that the peak PRL surge occurs in spite of active dopamine (DA) neurons, suggesting that it is generated by a nondopaminergic mechanism. Decreased TH activity in DA neurons in the SME and PP may prolong the PRL surge during the plateau phase, whereas increased DA activity coincides with the termination of the surge.

Somatostatin-containing neurons in the mouse brain: an immunohistochemical study and comparison with the rat brain.

The distribution of somatostatin-containing neurons in mice of both sexes was immunohistochemically examined and compared with that in rats. In radioimmunoassay the relative somatostatin content in the mouse brain was 2-3 times higher than that in the rat. The overall immunohistochemical staining for somatostatin was much stronger and more prominent in the mouse than in the rat. Although the distribution pattern of somatostatin immunoreactivity was basically the same between the two animals, several regions, especially the nucleus anterior hypothalami and the nucleus interpeduncularis, were found to contain large aggregates of somatostatin-immunoreactive neurons in the mouse brain but not in the rat. The electrolytic lesions to the nucleus anterior hypothalami caused a marked decrease in somatostatin immunoreactivity of the outer layer of the median eminence in the mouse. This suggests that the nucleus anterior hypothalami is an additional source of somatostatin for the median eminence in the mouse. The differences recognized between the species are interesting from functional and evolutionary points of view.

Topographic relations between tyrosine hydroxylase- and luteinizing hormone-releasing hormone-immunoreactive fibers in the median eminence of adult rats.

Employing electron microscopic double immunolabeling, we determined a close apposition of tyrosine hydroxylase (TH) and luteinizing hormone-releasing hormone (LHRH) nerve fibers in the rat median eminence (ME). These axo-axonic contacts occurred frequently in the internal and palisade zones, i.e. at the level of the fiber preterminals. In the superficial area of the ME, major TH fibers abutted on the basal lamina and some were projected into the pericapillary space of the portal vessels. Conversely, LHRH fibers were arrested by the endfeet of tanycytes in reaching the basal lamina.

Autoradiographic identification of prolactin binding sites in rat median eminence.

Binding sites for prolactin were localized and quantified in the female rat brain by in vitro autoradiographic analysis with iodine-125-labeled ovine prolactin. Following incubation, labeled prolactin bound preferentially to the median eminence and choroid plexus of the lateral and third ventricles. The addition of excess unlabeled ovine prolactin blocked binding of the labeled hormone in the choroid plexus, and attenuated prolactin binding in the median eminence. These results provide evidence for median eminence. These results provide evidence for prolactin-specific recognition sites in the median eminence, a region intimately involved in the hypothalamic regulation of prolactin secretion.

Alteration by thyroid hormones of TRH in the rat median eminence: role of the hypothalamic paraventricular nucleus.

We have shown that hypothyroidism produced a progressive decrease in the TRH content of the hypothalamic median eminence. In the present study, we explored whether thyroid hormones might alter TRH content in the median eminence by acting on the hypothalamic paraventricular nucleus (PVN) in the rat. After thyroidectomy, adult male rats received a hypothalamic deafferentation or electrolytic lesions in the bilateral PVNs. These rats were i.p. administered daily with T4 in a dose of 1.5 micrograms per 100 g body weight. Two weeks later, these rats were decapitated, and TRH in the median eminence and TSH in the peripheral blood were determined by RIAs. Thyroidectomy caused a significant reduction in TRH content of the median eminence. The hypothalamic deafferentation and PVN-lesions further reduced the TRH content. T4-replacement restored the decreased content of median eminence TRH to the normal level. The hypothalamic deafferentation and PVN-lesions prevented T4-induced restoration of TRH content. The present data imply that thyroid hormones regulate the amount of THR in the median eminence through acting on the hypothalamic PVN.

Influence of age on the control of thyrotropin secretion by thyrotropin-releasing hormone in the male rat.

Alterations with age in the control of thyrotropin (TSH) secretion by thyrotropin-releasing hormone (TRH) were evaluated at the hypothalamic and pituitary levels in young (3-5 months) and old (22-24 months) male rats. In the hypothalamus, TRH was quantified in the median eminence and in the mediobasal hypothalamus; in the adenohypophysis the membrane receptors for TRH were evaluated as well as the accumulation of TRH in the gland. As for TSH, its concentration was determined in the anterior pituitary gland and in plasma. In the hypothalamus, the concentration of TRH did not differ between young and old rats in the whole mediobasal hypothalamus, but it was significantly less in the old rats at the level of the median eminence (29.9 +/- 2.8 vs. 52.2 +/- 4.3 ng/mg protein). In the adenohypophysis, the density of receptors for TRH was greater in the old than in the young rats (23.2 +/- 3.2 vs. 13.7 +/- 1.1 fmol MeTRH/mg gland)--with no change in the affinity constant--, and the amount of TRH detected was larger (10.8 +/- 1.8 vs. 2.8 +/- 0.6 pg/mg gland), illustrative of an age-related increase in TRH accumulation in the pituitary gland. The latter results are contrasting with the findings of unchanged pituitary and plasma concentrations of TSH as well as unmodified TSH response to TRH in old rats. The present data concerning TRH and the analogy with previous observations regarding dopamine in old rats are indicative of reduced neuronal activities with age at the hypothalamic level associated with impairments in the processing of the hypothalamic hormones at the pituitary level.

Correlation between the stress-induced transient increase in corticotropin-releasing hormone content of the median eminence of the hypothalamus and adrenocorticotropic hormone secretion.

Intravenous angiotensin II and ether stress were found to produce a rapid, transient increase in the corticotropin-releasing hormone (CRH) content of the median eminence as measured by a radioimmunoassay employing an antibody against rat CRH(1-41). This confirms previous reports of transient increases in CRH measured by bioassay. The increase did not occur in the paraventricular region or in other parts of the brain. It occurred along with an increase in plasma adrenocorticotropic hormone (ACTH) when a second ether stress was administered 1 h after the first, and it also occurred when rats that had been adrenalectomized for 5 days were exposed to ether. The increases in CHR and the ACTH responses to ether were reduced or abolished by dexamethasone and pentobarbital. Four days after semicircular knife cuts in the posterior hypothalamus, resting CRH in the median eminence was increased but there was no further rise after ether stress. Plasma ACTH was normal at rest after the cuts, but the increase produced by ether was reduced. The ACTH responses to angiotensin II and immobilization were also reduced. Because the posterior knife cuts reduced hypothalamic catecholamine content, the effects of reducing hypothalamic norepinephrine and epinephrine by administration of the dopamine-beta-hydroxylase inhibitor diethyldithiocarbamate (DDC) were tested. Five hours after DDC, plasma ACTH was elevated but there was no further increase with ether stress. The median eminence CRH content was normal but failed to increase after exposure to ether.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of the central nucleus of the amygdala on the content of corticotropin-releasing factor in the median eminence.

The central nucleus of the amygdala possesses numerous neurons containing corticotropin-releasing factor (CRF). This study demonstrates a striking decrease of the CRF-like immunoreactivity in the median eminence at both 1 and 2 weeks after bilateral lesions of the amygdaloid central nucleus. Lesion of the amygdaloid central nucleus did not alter the neurophysin-like immunoreactivity in the internal zone of the median eminence, indicating the integrity of the efferent neurophysin-containing fibers of the supraoptic and paraventricular hypothalamic nuclei. However, there was a concomitant decrease of neurophysin and CRF-like immunoreactivity in the external zone of the median eminence. These results substantiate the hypothesis that the amygdaloid central nucleus can influence the content of CRF-like material in the median eminence via a multisynaptic pathway involving the synthesis of CRF at the level of the paraventricular nucleus of the hypothalamus. The exact mechanism by which lesion of the amygdaloid central nucleus influences the CRF content in the median eminence remains to be determined.