The central oxytocin pulse generator: a pacemaker for luteolysis.

Oxytocin (OT) is released from the neurohypophysis into the jugular vein of sheep in small 1-2 min pulses (ca. 10 pg/ml) in both cyclic and ovariectomized sheep. In intact cycling sheep, additional hour long bursts of OT (up to 200 pg/ml) occur in peripheral blood during luteolysis at intervals of 6 to 9 hrs which appear to regulate large luteolytic pulses of uterine prostaglandin F2a (PGF2a). Since the ovine corpus luteum (CL) also synthesizes OT, experiments were performed to distinguish between the relative contributions of the neurohypophysis and the CL to the large bursts of OT secreted during luteolysis. Two models were used. First, ovariectomized sheep were given exogenous E and/or P by constant infusion to simulate levels during the estrous cycle. Second, in tact cycling sheep, the CL was surgically excised during the luteal phase to exclude the CL as a source of OT and, at the same time, subject the animals to the withdrawal of P. Pulses of OT in jugular vein plasma were determined by RIA or biometry of the uterus. The findings are summarized as follows: In ovariectomized sheep, maintained on low E (0.05 g/hr) to preserve the OT pulse generator, infusion of E (1 microgram, 2 micrograms or 4 micrograms/hr) for 12 to 36 hr, caused a series (4 to 6) of rapid increases in OT pulse frequency each lasting 1 to 2 hrs at intervals of 3 hrs. The time of onset of high frequency pulses was dose-dependent. Withdrawal of 10 day infusions of P (500 micrograms/hr) superimposed on low E (0.05 microgram/hr) also evoked a series of high frequency episodes of OT pulses beginning 24 hrs after P withdrawal. In intact sheep, surgical removal of the CL resulted in a series of high frequency pulses similar in duration and frequency to those following the withdrawal of P in the ovariectomized animal. We conclude that: (1) an increase in E or returning E action causes the OT pulse generator to alter its frequency intermittently thus producing a series of 4 to 6 episodes of high frequency pulses of OT. (2) Similar changes can be evoked by withdrawal of P either by terminating an infusion of P in the presence of E in the ovariectomized sheep or by surgically removing the CL from the ovary in the intact sheep. (3) At the end of the reproductive cycle, the central OT pulse generator appears to act as a pacemaker which, acting on the endometrial OT receptors, triggers a series of pulses of PGF2a from the uterus and hence causes regression of the CL. In the sheep and other ruminants, an intermittent supplemental secretion of OT from the CL, triggered via the central OT pulse generator, may also be required to amplify the luteolytic pulses of PGF2a from the uterus. (4) In addition to the well established interaction of ovarian steroid hormones, and the hypothalamic pituitary system for the initiation of the reproductive cycle via the gonadotrophins, there is now good evidence for an interaction of ovarian steroids and the posterior pituitary for terminating the reproductive cycle.

Vasopressin transport regulation is coupled to the synthesis rate.

Vasopressin is synthesized in the perikarya of magnocellular neurons and is transported down long axons to the storage terminals of the posterior pituitary. To maintain stable pituitary stores following vasopressin secretion, the hypothalamus must synthesize and transport an amount of new vasopressin, equivalent to the amount released. Vasopressin release and synthesis rate can be chronically upregulated or suppressed relative to basal levels, depending on the demand for vasopressin. We studied whether vasopressin transport was similarly regulated during situations of varying demand. During chronic hyponatremia, when synthesis of vasopressin was reduced to undetectable levels, transport of vasopressin was also markedly decreased, as evidenced by continued presence of vasopressin in the transport system. Upregulation of transport was demonstrated by measuring pituitary accumulation of vasopressin in rats whose pituitary stores were initially depleted by hypernatremia and in whom subsequent release was suppressed by hyponatremia. In hypernatremic rats, transport of vasopressin was increased fivefold over baseline as determined by pituitary accumulation, and this elevated rate persisted for 7 days in the absence of release. This study demonstrates that axonal transport of vasopressin is a regulated process and is linked to synthesis rate rather than release.

Quantitation of changes in the content of neurohypophyseal peptides in hypothalamic nuclei after adrenalectomy.

In adrenal insufficiency there is an increase in the content of vasopressin in the external zone of the median eminence as determined by immunohistology. We studied rats after bilateral adrenalectomy and obtained punched samples of the supraoptic, the paraventricular, and the suprachiasmatic nuclei, and from the median eminence and the posterior pituitary. Vasopressin, neurophysin and oxytocin were all decreased in content in the posterior pituitary but in the suprachiasmatic nucleus and in the median eminence there was a significant increase in the content of vasopressin. The increased levels of vasopressin in the median eminence are interpreted as representative of the content in axons and demonstrate a large increase of transport of vasopressin. In the SON and PVN there was no increase in the content of vasopressin during this state of augmented synthesis and transport, indicating that vasopressin moves rapidly from the site of synthesis into the transport system. The increased content of vasopressin in the suprachiasmatic nucleus shows that vasopressin at this level also varies with changes in adrenal function. Levels of oxytocin throughout the neurohypophysis did not change in parallel with vasopressin demonstrating relatively selective responses of the neurohypophyseal hormones.

Vasopressin-dependent and -independent control of the release of adrenocorticotropin.

We tested the possibility that vasopressin mediates the responses of adrenocorticotropin (ACTH) to electrical stimulation of various areas of the hypothalamus. Thirty-three cats were anesthetized with chloralose-urethane, immobilized with gallamine, and respired artificially. Plasma ACTH was measured by RIA. Intraventricular administration of antiserum to vasopressin blocked the release of ACTH induced by electrical stimulation of the paraventricular nucleus (PVN), suggesting a role for the vasopressinergic projection from PVN to the external zone of the median eminence. In contrast, the release of ACTH induced by stimulation of areas ventral to PVN was unaffected by the antiserum. Thus, there is at least one corticotropin releasing factor released from nuclei other than PVN that is distinct from vasopressin.

Control of release of adrenocorticotropin and vasopressin by the supraoptic and paraventricular nuclei.

To determine the relative roles of the hypothalamic supraoptic (SON) and paraventricular (PVN) nuclei in the control of the release of vasopressin and of ACTH, we have examined the hormonal responses to electrical stimulation (200 microA, 0.2 msec, 100 Hz, 20 sec) of these regions. Cats were anesthetized with chloralose-urethane. Blood samples were taken 30 sec before stimulation and 1.5 min poststimulation. ACTH and vasopressin were measured by RIA. Electrical stimulation of the caudal pole of the SON increased vasopressin in plasma (1.82 +/- 0.41 microU/ml, n = 17, P less than 0.01) and decreased ACTH (-26 +/- 4 pg/ml, n = 13, P less than 0.01). In contrast, stimulation of the PVN increased vasopressin (2.01 +/- 0.60 microU/ml, n = 7, P less than 0.001) and increased ACTH (107 +/- 20 pg/ml, n = 32, P less than 0.01). Previous work has shown that vasopressinergic neurons of PVN, but not of SON, project to the zona externa of the median eminence. Other have suggested that the retrograde flow of blood from the neural lobe to the median eminence and thence to the anterior lobe would allow vasopressin to influence the release of ACTH. The present results indicate that both SON and PVN facilitate the release of vasopressin. However, PVN facilitates, but SON inhibits the release of ACTH. These findings suggest that the projection from PVN to the zona externa of the median eminence mediates the release of ACTH and that retrograde flow from the neural lobe is not important in the control of ACTH release during modest and transient increases in the release of vasopressin.

Levels of neurohypophyseal peptides in the rat during the first month of life. I. Basal levels in plasma, pituitary, and hypothalamus.

Vasopressin, oxytocin, and neurophysin were measured by RIA in the pituitary, hypothalamus, and plasma (except oxytocin) of the rat during the first month of life. In plasma, vasopressin was less than 1.7 microU/ml in most animals. Neurophysin was elevated above adult levles on day 2 and decreased with age. The three peptides were present in the pituitary at birth, but in amounts less than 1% of the adult level. The vasopressin content of the pituitary increased rapidly in the first days after birth, while the levels of oxytocin and neurophysin remained low until 8 days and then increased between 8-21 days. The ratio of vasopressin to oxytocin in the pituitary was 4.4 at birth and reached unity (the ratio in the adult) at 30 days. At birth, the moles of neurophysin in the pituitary relative to the moles of hormone (oxytocin plus vasopressin) was low (0.15), largely due to a molar excess of vasopressin. The ratio of neurophysin to hormone reached unity at 21-30 days. Assays to detect vasotocin gave negative results. It is postulated that a precursor neurophysin which was related to vasopressin was present in the fetal rat but was not measured in our study.

Levels of neurohypophyseal peptides in the rat during the first month of life. II. Response to physiological stimuli.

Levels of vasopressin, oxytocin, and neurophysin were measured by RIA in the hypothalamus, pituitary, and plasma of infant rats (2-30 days old). At all ages, ip injection of a hypertonic solution of 5 g/100 ml NaCl produced a marked increase in levels of vasopressin and neurophpysin in plasma, up to 21 microU/ml and 51 ng/ml, respectively. After dehydration, there was a decrease of 26-38% in the levels of neurohypophyseal peptides in the pituitary. Depletion of neurohypophyseal peptides from the pituitary was greater after 24 h of dehydration in younger rats (26%) than in older rats (7%). Levels of vasopressin in plasma were less than 1.7 microU/ml after dehydration in younger rats but were greater in older rats. Immaturity of the neurohypophysis may contribute to the inability of newborn rats to withstand prolonged dehydration.

Plasma neurophysin and vasopressin in the rat: response to adrenalectomy and steroid replacement.

Impaired excretion of a water load is known to occur in adrenal insufficiency and to be corrected by administration of glucocorticoid. Such impairment has been related to either a loss of a permissive effect of glucocorticoids on the diluting segments of the nephron or to an alteration of release, turnover, or action of antidiuretic hormone. Specific and sensitive RIAs for arginine vasopressin and neurophysin were utilized to measure plasma and pituitary levels of neurohypophyseal peptides at baseline and after an intragastrically administered water load. Conscious, unanesthetized, and nonstressed sham-operated, adrenalectomized, and adrenalectomized prednisone-treated rats were studied. The results demonstrate a significant elevation in vasopressin and neurophysin in plasma in adrenalectomized rats maintained in a normal state of hydration. After water loading, the adrenalectomized rats diluted their plasma osmolality but had a decreased urinary volume, increased urinary osmolality, and elevated vasopressin and neurophysin in their plasma. In the pituitary, vasopressin and neurophysin were depleted in adrenalectomized rats, indicating increased secretion of these peptides. It is concluded that elevated vasopressin in plasma may be an important factor in the incomplete water diuresis in adrenal insufficiency.

Plasma neurophysin levels in monkeys: emphasis on the hypothalamic response to estrogen and ovarian events.

Specific radioimmunoassays for human neurophysins released in response to estrogen (estrogen-stimulated neurophysin, ESN) and nicotine (nicotine-stimulated neurophysin, NSN) have been used to measure two similar neurophysins in rhesus monkey plasma. As in the human, concentrations of rhesus monkey neurophysins in plasma were specifically produced a marked increase of plasma NSN concentrations in the monkey. Estradiol benzoate administered intramuscularly consistently produced an increase in plasma ESN concentrations in normal cycling and castrate monkeys. ESN response to estrogen was exclusively positive and occurred approximately 10 hours after an injection of estradiol benzoate intramuscularly. Plasma samples obtained throughout the mid-cycle were measured and a characteristic rise in estrogen and LH, and a more prolonged rise in ESN were found. Our data indicate that the ESN and LH responses to estrogen stimulation are temporally related events and that the assay of ESN in plasma may be of unique value as it directly reflects the hypothalamic response to changes in estrogen secretion.