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H. M. A. MEIJS-ROELOFS
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SUMMARY

Electrical stimulation of the hypothalamus with biphasic pulses was performed in immature female rats. When performed at 27 days of age or later, electrical stimulation in the arcuate nucleus region advanced puberty in all animals, as did stimulation of the anterior hypothalamus at 29 days of age or later. Stimulation in younger rats did not uniformly advance puberty. The responsiveness to electrical stimulation thus seems to develop a few days earlier in the arcuate nucleus region than in the anterior hypothalamus.

In a second experiment the possible involvement of follicle-stimulating hormone (FSH) in the advancement of puberty was investigated: the simplified augmented ovarian weight assay for endogenous FSH was performed in rats stimulated in the arcuate nucleus region as well as in controls. A marked increase in ovarian weight, indicating increased FSH levels, was demonstrated in all animals stimulated on day 27 or later; at earlier ages only a percentage of the stimulated animals responded. This percentage paralleled the percentage of animals that showed advancement of puberty.

It is concluded that electrical stimulation in both the arcuate nucleus region and the anterior hypothalamus advances the onset of puberty. It is suggested that electrical stimulation causes increased plasma FSH levels and, in consequence, precocious puberty.

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JEAN E. M. BUTLER
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B. T. DONOVAN
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Surgical isolation of the hypothalamus from the rest of the brain was used in a study of the hypothalamic control of gonadotrophin secretion. Rats with hypothalamic islands of differing sizes and animals in which the hypothalamus was partially deafferentated were prepared. Oestrous cycles were followed by taking daily vaginal smears and the reproductive state assessed from histological examination of the genital tract collected at autopsy 28 days postoperatively. In rats with 'large' and 'long' hypothalamic islands persistent vaginal oestrus ensued and the ovaries were follicular. Rats in which smaller islands disconnected the suprachiasmatic nuclei from the median eminence displayed persistent vaginal dioestrus and the ovaries contained corpora lutea. Rats with frontal cuts placed anterior to the suprachiasmatic nuclei behaved similarly to rats with 'large' and 'long' islands in developing persistent vaginal oestrus, but when the anterior connexions to the hypothalamus were severed caudal to the suprachiasmatic nuclei the animals had oestrous cycles which were sometimes interrupted by periods of dioestrus of pseudopregnancy length. Posterior and transverse cuts in the hypothalamus did not alter the oestrous cycle although there was some indication that luteal function was favoured.

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J. Alaghband-Zadeh
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S. Fenton
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K. Hancock
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J. Millettt
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H. E. de Wardener
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Acetone extracts from a variety of rat tissues were tested for their ability to stimulate renal glucose-6-phosphate dehydrogenase (G6PD) activity at 2 min in an in-vitro cytochemical assay which is a marker of the sodium potassium-dependent adenosine triphosphatase (Na+-K+-ATPase) inhibiting activity. Extracts of the hypothalamus were the only ones found to be active in this system. Acetone extract of hypothalamus also inhibited renal Na+-K+-ATPase activity in vitro. The G6PD-stimulating activity from one hypothalamus was about 10000 to 100 000 times greater than that of 1 ml plasma. The G6PD-stimulating activity of hypothalamic extracts from rats which had been on a high sodium intake for 4 weeks were approximately 150 times more active than those obtained from rats which had been on a low sodium diet. The G6PD-stimulating activity of the corresponding plasma was sixfold more active. These findings suggest that a circulating sodium transport inhibitor(s) may be secreted from the hypothalamus.

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R. E. PETER
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The effect on thyroid activity of a systemically ineffective dose of thyroxine (T4) implanted in the hypothalamus or pituitary of goldfish was tested. Thyroid activity was decreased by T4 implantation in either location, indicating that T4 has a negative feedback effect on the pituitary causing a decrease in thyrotrophin secretion, and a positive feedback effect on the hypothalamus stimulating the secretion of thyrotrophin inhibitory factor (TIF). Fish with a T4 or blank-control implant in the pituitary that had a damaged pituitary stalk, as a result of the operative procedures, were hyperthyroid, suggesting either that TIF is more effective in suppressing thyrotroph activity than T4 and that the effect of T4 was masked by the absence of TIF, or, less likely, that T4 negative feedback in the pituitary is not effective independent of TIF. The results were compared with the information about T4 feedback in mammals.

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L. ORSI
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J. H. DENARI
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C. A. NAGLE
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D. P. CARDINALI
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J. M. ROSNER
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After decades of contradictory results it has become apparent that the mammalian pineal exerts an inhibitory influence on several neuroendocrine functions, namely the hypophysial-gonadal system, the adrenal cortex and the thyroid gland (Wurtman, Axelrod & Kelly, 1968). Among pineal secretory products the best characterized is melatonin (5-methoxy-N-acetyltryptamine), which reverses many of the endocrine changes that follow pinealectomy and mimics the effect of injecting pineal extracts into experimental animals. In the last 15 years considerable information has been accumulated on the mechanisms involved in the regulation of pineal melatonin synthesis; very little is known, however, about the biochemical processes evoked by melatonin in its target organs. Exogenous melatonin has been shown to become concentrated within the hypothalamus (Antón-Tay & Wurtman, 1969; Cardinali, Hyyppä & Wurtman, 1973). Implants of melatonin in the hypothalamus decrease pituitary gonadotrophin levels (Fraschini, 1969); moreover, melatonin perfusions of the third ventricle decrease plasma levels of

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K. SEIKI
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M. MIYAMOTO
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A. YAMASHITA
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M. KOTANI
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In a previous paper (Seiki, Higashida, Imanishi, Miyamoto, Kitagawa & Kotani, 1968), it was briefly reported that specific progesterone-binding receptors probably do not exist in the hypothalamus and pituitary gland, because these sites did not show selective uptake of progesterone compared with other parts of the brain after i.v. administration of [3H]progesterone to rats ovariectomized in dioestrus 2 weeks before. Other workers (Watanabe, Saha & Layne, 1968) have failed to show any selective uptake of tritiated norethindrone by the hypothalamus and pituitary gland. To confirm these results, two further experiments have been carried out as follows.

Female Wistar rats (about 200 g. body weight) exhibiting 4-day oestrous cycles, as determined by vaginal smears, were used. In expt 1, the animals in four stages of the oestrous cycle were each given an i.v. injection of 30 μc (1·7 × 10−3 μmole) [7α-3H]progesterone (sp. act. 18·1 c/μmole)in 0·2

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D. T. DAVIES
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B. K. FOLLETT
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SUMMARY

Experiments were undertaken to localize those hypothalamic areas in the male quail (Coturnix coturnix japonica) where electrical stimulation would increase LH secretion. The posterior basal hypothalamus was stimulated with rectangular pulses (height 500 μA) through a bipolar electrode for 6 min, blood samples being taken for LH assay 20 min before, and 2, 10, 20 and 30 min after stimulation. The highest plasma concentration was observed in the 2 min sample. Over the next 30 min the LH level decreased to the resting concentration. The relative increase in LH level was greatest in sexually immature quail and least in photostimulated castrated birds, although the highest absolute levels were seen in the castrated quail. There were no statistical differences between the magnitude of the LH increases in sexually immature, mature and castrated quail.

Various hypothalamic regions were then stimulated with a smaller current (200 μA) applied for only 2 min. A highly significant rise in LH followed stimulation of either the tuberal hypothalamus (postero-dorsal part of the infundibular nuclear complex, PD-INC), or the preoptic region (POR) while stimulation 0·5–1·5 mm away from these regions did not change LH secretion. Stimulation of the anterior basal hypothalamus, or of the suprachiasmatic area, caused a significant rise in LH concentration although this was less than that seen after stimulation of the POR. Stimulation in the POR or the PD-INC was ineffective if the tuberal hypothalamus had been deafferentated surgically some days previously. The data complement the studies in which destruction of the PD-INC or the POR by electrolytic lesions has been shown to block photoperiodically induced testicular growth and LH secretion.

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B. A. CROSS
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1. Electrical stimulation of the supraoptico-hypophysial system gave rise to milk ejection from cannulated teats in anaesthetized lactating rabbits. Stimuli applied in the vicinity of the paraventricular nucleus produced signs of sympathetic activity and no milk-ejection responses in the intact animal, but after acute adrenalectomy milk-ejection responses could be elicited from this region.

2. Stimulation of the dorsal, lateral and posterior areas of the hypothalamus, but not of the ventral tuberal regions, caused pupillary dilatation, exophthalmos and hyperpnoea, and inhibited the milk-ejection response to oxytocin injected intravenously 10–15 sec after the end of the stimulus. The inhibitory effect could be simulated by injection of 1–5 μg adrenaline. It was abolished by bilateral adrenalectomy.

3. Inhibition of the milk-ejection response to oxytocin was also produced by stimulation of the splanchnic nerve supply to the adrenal glands, and of the sympathetic nerve supply to the mammary glands. After bilateral adrenalectomy some inhibitory effect on milk-ejection was apparent when oxytocin was injected during prolonged stimulation of the sympathetic centres of the hypothalamus.

4. Adrenaline was 1½ to 4 times more active in blocking the milk-ejection response to intravenous oxytocin than noradrenaline.

5. Stimulation of the hypothalamus or mammary sympathetic nerves, and injection of adrenaline or noradrenaline did not inhibit the milk-ejection response to mechanical stimuli applied direct to the mammary glands.

6. The inhibition of the milk-ejection response to oxytocin produced by stimulation of the hypothalamus was associated with inability of the young to remove milk from the mammary glands during suckling.

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P. E. BELCHETZ
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GLENYS GREDLEY
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DAPHNE BIRD
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R. L. HIMSWORTH
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Bilateral injections of tri-iodothyronine (T3, 2 ng in 2 μl artificial cerebrospinal fluid, CSF) were made into the hypothalami of 15 hypothyroid rhesus monkeys (Macaca mulatta) prepared with chronically implanted intrahypothalamic cannulae. In 29 out of 96 such injections, the concentration of thyrotrophin (TSH) in the plasma fell rapidly by more than 30% and returned to the basal value over the succeeding 48 h, in 13 experiments the fall was 20–30% and in the remaining 54 experiments the level of TSH was unaltered. With [125I]T3, the extent of diffusion of T3 through the hypothalamus in these experiments was shown to be very limited. The hypothalamic injection sites were subsequently identified histologically and it was found that in those experiments in which the level of TSH in the plasma had fallen, T3 had been injected into either the dorsomedial nucleus or the lateral hypothalamic area extending into the preoptic region. Injection of artificial CSF alone into these areas of the hypothalamus did not affect the concentration of TSH. Direct intrapituitary injections of T3 (4 ng) resulted in small and inconstant changes in the concentration of TSH in the plasma and injections of T3 (400 ng) into the third ventricle were without any effect. These experiments demonstrate that T3 can rapidly inhibit the secretion of TSH by a direct action upon defined parts of the hypothalamus.

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D. G. MONTEMURRO
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SUMMARY

The ventral hypothalamus and the pituitary gland of the rat were examined with the light microscope in sections fixed in an osmium tetroxide solution without further staining. The histology of the neurohypophysis could be observed with remarkable clarity. Herring bodies and axons of the supra-optico-neurohypophysial tract show as grey homogeneous structures. Pituicytes appear as round or variously shaped cells whose cytoplasm is crowded with osmiophilic granules. The granules vary in size from 0·5 to about 2·5 μ, and present either as solid or as ring-shaped black structures. They stained neither with aldehyde-thionine, nor with chrome-alum haematoxylin, nor with the PA-Schiff techniques but stained with Sudan black B. They were found in profusion in the median eminence of the hypothalamus and the infundibular stem in an apparent extracellular location surrounding fibre bundles and capillaries of the hypophysial portal plexus. Two types of cells were identifiable in the supraoptic nucleus of the hypothalamus. Smaller, angular neurones stained densely with osmium and with aldehyde-thionine and larger, more rounded neurones stained only lightly with osmium and did not stain with aldehyde-thionine. A peculiar laminated arrangement of structures in the infundibular recess in the region of the arcuate nucleus was noted.

Several distinct types of cells in the adenohypophysis were identifiable on morphological grounds: thyrotrophic basophils, gonadotrophic basophils, acidophils and chromophobes. The possibility of a neuroendocrine role of the pituicytes is discussed.

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