Paradoxical elevation of growth hormone by intraventricular somatostatin: possible ultrashort-loop feedback.

Somatostatin, the growth hormone-inhibiting factor, when microinjected into the third ventricle of the rat brain, paradoxically induced the release of growth hormone. A pituitary site of action having been ruled out, this result supports the concept that exogenous somatostatin within the hypothalamus acts either to suppress the release of somatostatin from somatostatin-containing neurons, possibly via an ultrashort-loop feedback mechanism, or to augment release of hypothalamic growth hormone-releasing factor, thereby inducing a release of growth hormone. Injection of somatostatin into the third ventricle also decreased plasma concentrations of luteinizing hormone, follicle-stimulating hormone, and thyroid-stimulating hormone, probably by inhibiting the release of luteinizing hormone-releasing factor and thyrotropin-releasing factor.

Arginine vasopressin as a thyrotropin-releasing hormone.

Although hypothyroidism (with concomitant increased levels of thyroid-stimulating hormone) has been associated with elevated plasma vasopressin, the role that vasopressin plays in controlling thyroid-stimulating hormone secretion from the adenohypophysis is not understood. In two in vitro pituitary cell systems, vasopressin caused a specific and dose-related release of thyroid-stimulating hormone from cells that was equal in potency to that elicited by thyrotropin-releasing hormone, the primary acknowledged regulator of thyroid-stimulating hormone release. When injected into the hypothalamus, however, vasopressin specifically inhibited the release of thyroid-stimulating hormone. Thus, vasopressin may exert differential regulatory effects on thyroid-stimulating hormone secretion in the hypothalamus and pituitary gland.

Adaptive Biomedical Innovation: Evolving Our Global System to Sustainably and Safely Bring New Medicines to Patients in Need.

The current system of biomedical innovation is unable to keep pace with scientific advancements. We propose to address this gap by reengineering innovation processes to accelerate reliable delivery of products that address unmet medical needs. Adaptive biomedical innovation (ABI) provides an integrative, strategic approach for process innovation. Although the term "ABI" is new, it encompasses fragmented "tools" that have been developed across the global pharmaceutical industry, and could accelerate the evolution of the system through more coordinated application. ABI involves bringing stakeholders together to set shared objectives, foster trust, structure decision-making, and manage expectations through rapid-cycle feedback loops that maximize product knowledge and reduce uncertainty in a continuous, adaptive, and sustainable learning healthcare system. Adaptive decision-making, a core element of ABI, provides a framework for structuring decision-making designed to manage two types of uncertainty - the maturity of scientific and clinical knowledge, and the behaviors of other critical stakeholders.

"Threshold-crossing": A Useful Way to Establish the Counterfactual in Clinical Trials?

A central question in the assessment of benefit/harm of new treatments is: how does the average outcome on the new treatment (the factual) compare to the average outcome had patients received no treatment or a different treatment known to be effective (the counterfactual)? Randomized controlled trials (RCTs) are the standard for comparing the factual with the counterfactual. Recent developments necessitate and enable a new way of determining the counterfactual for some new medicines. For select situations, we propose a new framework for evidence generation, which we call "threshold-crossing." This framework leverages the wealth of information that is becoming available from completed RCTs and from real world data sources. Relying on formalized procedures, information gleaned from these data is used to estimate the counterfactual, enabling efficacy assessment of new drugs. We propose future (research) activities to enable "threshold-crossing" for carefully selected products and indications in which RCTs are not feasible.

From adaptive licensing to adaptive pathways: delivering a flexible life-span approach to bring new drugs to patients.

The concept of adaptive licensing (AL) has met with considerable interest. Yet some remain skeptical about its feasibility. Others argue that the focus and name of AL should be broadened. Against this background of ongoing debate, we examine the environmental changes that will likely make adaptive pathways the preferred approach in the future. The key drivers include: growing patient demand for timely access to promising therapies, emerging science leading to fragmentation of treatment populations, rising payer influence on product accessibility, and pressure on pharma/investors to ensure sustainability of drug development. We also discuss a number of environmental changes that will enable an adaptive paradigm. A life-span approach to bringing innovation to patients is expected to help address the perceived access vs. evidence trade-off, help de-risk drug development, and lead to better outcomes for patients.

Blockade of growth hormone-releasing factor (GRF) activity in the pituitary and hypothalamus of the conscious rat with a peptidic GRF antagonist.

Microinjection of synthetic GRF into the cerebroventricles or hypothalamus of the rat produces a number of neural effects, including the suppression of GH secretion, possibly representing a negative ultrashort loop autoregulation of GRF and/or stimulation of somatostatin neurosecretion. To demonstrate that such neuromodulation acts physiologically through endogenous GRF activity, the peptidic GRF antagonist (N-Ac-Tyr1,D-Arg2)GRF-(1-29)-NH2 was used to block the action of GRF on its presumed receptors in the hypothalamus. First, to establish the efficacy of the antagonist to block GRF receptors in the anterior pituitary, we injected the antagonist iv at doses of 2, 20, and 50 micrograms or saline (controls) into conscious male rats fitted with jugular cannulae. Sequential blood sampling every 15 min for 6 h between 1000-1600 h showed that 50 micrograms antagonist, iv, significantly suppressed the two periods of spontaneous release of radioimmunoassayable GH in controls in the morning and afternoon. A dose of 20 micrograms, iv, lowered mean plasma GH between 1400-1500 h (P less than 0.025), while the 2-microgram dose was without effect. The GRF antagonist was then microinjected into the third ventricle (3V) of conscious male rats at doses of 0.5 and 8.0 ng in 2 microliter sterile saline. The 8.0-ng dose of 3V antagonist elicited a 3-fold increase in the morning peak of GH (nanograms per ml): 3V antagonist, 159.0 +/- 62.0; 3V control, 51.0 +/- 21.9 (P less than 0.05). The 0.5-ng dose was without effect. Finally, we observed that pretreatment with the GRF antagonist 3V (10 ng), followed 15 min later by 10 ng rat GRF administered 3V, completely blocked the GRF-induced suppression of pulsatile GH release observed earlier. Both the systemic and central effects of the antagonist were specific to the control of GH, since PRL concentrations were unaltered. These results 1) have demonstrated the ability of a peptidic GRF antagonist to specifically suppress pulsatile GH release after its systemic administration, presumably by acting on pituitary GRF receptors, and 2) support the notion that GRF receptors are also present in the hypothalamus and are available for the physiological mediation of GRF-induced inhibition of GH release by a central mechanism.

Effect of destruction of the dorsal anterior hypothalamus on follicle-stimulating hormone secretion in the rat.

The role of the paraventricular nucleus-dorsal anterior hypothalamus (PVN-DAHA) in the control of anterior pituitary gland secretion of FSH and LH in castrated male and female rats was examined. Bilateral radiofrequency lesions of the PVN-DAHA in chronically ovariectomized (OVX) rats lowered plasma FSH levels by 33% (P less than 0.005) compared to values in unoperated and sham-operated control rats; plasma LH concentrations were unaltered. RIA of median eminence (ME) LHRH concentrations in these animals revealed no differences among the three experimental groups. Other categories of diencephalic destruction did not result in this pattern of selectively reduced FSH release. Bilateral radiofrequency destruction of the PVN-DAHA also attenuated by 50% (P less than 0.025 to P less than 0.005) the progesterone-induced surge of FSH in estrogen-primed OVX rats. Progesterone-induced LH release was unaffected by PVN-DAHA lesions. Other lesion categories failed to show the same result. Bilateral ablation of the PVN-DAHA in male rats resulted in a selective diminution of the postcastration rise of plasma FSH beginning 48 h postcastration (P less than 0.05 to P less than 0.005) and persisting for 14 days (P less than 0.005) after orchidectomy, thus revealing the time course and permanence of this procedure on plasma FSH levels. The postcastration rise of plasma LH levels was not affected by PVN-DAHA lesions. The concentration of ME LHRH was the same among orchidectomized male rats whether they bore PVN-DAHA lesions, sham lesions, or no lesions. In summary, destruction of the PVN-DAHA was found to reduce significantly the elevation of plasma FSH, but not LH, in the OVX rat and the estrogen-progesterone-stimulated OVX rat. PVN-DAHA lesions also attenuated the postcastration rise of FSH, but not that of LH, in the male. The failure of lesions of the PVN-DAHA to alter ME LHRH concentrations in the face of decreased FSH release does not prove that LHRH release is totally unaffected by this procedure. This finding is, however, consistent with the concept that diminished FSH secretion could be the result of a deficiency of a hypothalamic releasing factor (FSH-releasing factor?) other than that of the LHRH decapeptide.

Inhibition of pulsatile growth hormone (GH) secretion and somatic growth in immature rats with a synthetic GH-releasing factor antagonist.

We previously reported that systemic administration of the recently described GRF peptide antagonist (N-Ac-Tyr1,D-Arg2)GRF-(1-29)-NH2 to adult male rats would suppress the pulsatile release of GH. In the present study, we have sought to determine whether this same antagonist would be efficacious in immature male rats to block spontaneous GH secretion and, as a result, retard several parameters of somatic growth. Indwelling Silastic catheters were placed into the jugular veins of immature male rats (120-140 g) at 29 days of age. After a recovery period of 48 h, beginning at 1000 h, 100-400 micrograms/kg GRF antagonist or its vehicle (controls) were injected iv immediately after withdrawing an initial blood sample from conscious undisturbed animals. Subsequent samples were obtained every 20 min until 1520 h. Red blood cells were resuspended in a restorative volume of saline and reinjected after each blood sample. Results showed that both doses of antagonist prevented the two major periods of episodic GH release observed in controls. For example, mean plasma GH (+/- SEM; nanograms per ml) at 1120 h was 9.0 +/- 2.7 in antagonist-treated rats and 37.1 +/- 5.1 in controls (P less than 0.05). Mean plasma GH (+/- SEM) at 1340 h was 10.8 +/- 3.7 in antagonist-treated rats and 38.8 +/- 9.6 in controls (P less than 0.05). Injection of 400 micrograms/kg of the structurally related VIP antagonist (N-Ac-Tyr1,D-Phe2)GRF-(1-29)-NH2, iv failed to suppress spontaneous GH release. GRF antagonist (100 micrograms/kg) was next administered twice daily iv for 4 days to 31-day-old rats in metabolic cages. This treatment essentially arrested the normal rapid body weight gain, significantly suppressed increases in body and tail lengths, and reduced increases in heart and kidney weights (P less than 0.01). Food intake and fecal output were unchanged by antagonist treatment and, therefore, did not contribute to the observed effects. These results support the idea that a number of tissues and organs are stimulated by the pulsatile secretion of GH and that a peptidic GRF receptor antagonist is useful in blocking episodic GH release in immature animals. As a consequence, this specific antagonist is effective in suppressing numerous aspects of somatic growth.

Pulsatile release of follicle-stimulating hormone in ovariectomized rats is inhibited by porcine follicular fluid (inhibin).

While the pulsatile release of LH in ovariectomized animals has been well established, little information exists concerning the pulsatile release of FSH and the factors involved in its regulation. The present studies analyzed the characteristics of episodic FSH secretion and examined the effects of ovarian feedback signals on that pattern. From conscious unrestrained ovariectomized rats, blood was collected at 10-min intervals for a period of 3 h. A pulsatile pattern of FSH was observed in the plasma of control rats, with an average frequency of 4 peaks/3, h, while LH oscillations occurred at an average of almost 6 peaks per 3 h. The injection of porcine follicular fluid (PFF; 0.5 ml), a source of inhibin, reduced FSH peak frequency to 2.5 peaks/3 h (P less than 0.005). In the third hour of blood sampling, PFF also significantly lowered the average FSH peak amplitude (P less than 0.05), the trough value, and the average mean value for that hour. In contrast, none of these parameters for LH was altered by PFF. Estradiol benzoate (EB; 20 micrograms, sc, 24 h before bleeding) significantly reduced LH peak frequency, mean trough value, and average mean value, whereas it only reduced the mean FSH average value and mean trough value. Combined treatment with EB and PFF synergized to decrease the mean value, trough value, peak frequency, and peak amplitude of episodic LH release. Pituitary responsiveness to LHRH was evaluated after the treatments described above. PFF inhibited the response of FSH to LHRH, but not that of LH. EB by itself potentiated the release of both hormones. The combined EB/PFF treatment resulted in a decreased pituitary response in terms of FSH and LH compared to that of the EB-treated control group. We conclude that the release of FSH occurs in a pulsatile fashion and, further, that this pattern of release is regulated by PFF (inhibin). It was also shown that an intriguing synergism between estradiol and inhibin exists to suppress LH release.

Evidence for a hypothalamic site of action of inhibin to suppress FSH release.

To determine if inhibin has a hypothalamic site of action to suppress FSH release, highly purified inhibin preparations from the rete testis fluid (RTF) of rams were injected into adult male rats which had been orchidectomized (ORDX) 24 hours earlier. Third ventricular (3rd V) injection of a potent inhibin fraction (RTF 38-I) significantly depressed plasma FSH concentrations, without influencing LH, 4-10 h after treatment. A less active preparation of inhibin (RTF 38-II) at the same dose had no effect. A higher dose of another less potent fraction (RTF 1A) significantly reduced FSH 2-6 h following 3rd V administration, accompanied by slight but significant decrements in LH at 2 and 4 only. To determine responsiveness of the pituitary, luteinizing hormone-releasing hormone (LHRH) was injected intravenously at 6 h. It induced similar elevations of FSH and LH in inhibin- and saline-treated groups. Systemic administration of RTF 38-II at a dose 2.5-fold higher than the dose effective centrally failed to modify either FSH or LH levels up to 6 h. These results provide evidence that inhibin from the male can preferentially suppress FSH release by a CNS site of action in addition to its well-known pituitary site of action.

Effects of intraventricular growth hormone-releasing factor on growth hormone release: further evidence for ultrashort loop feedback.

We examined the effects of cerebroventricular injection of synthetic human GH-releasing factor [hGRF-(1-44)] on regulation of GH release in conscious male rats. These results were compared with the direct effects of hGRF on hormone released from dispersed anterior pituitary cells. Administration of two higher doses of hGRF (200 and 2000 ng) into the third ventricle (3V) produced a dose-related increase in plasma GH levels (P less than 0.001). Injection of hGRF into the 3V at two lower doses actually reduced GH release. Infusion of 20 ng (5 pmol) hGRF reduced plasma GH from 5-60 min (P less than 0.005), with a maximum suppression of 66%. The 2-ng (0.5-pmol) dose decreased GH secretion by 45% (P less than 0.05). hGRF stimulated a significant and dose-dependent release of GH from dispersed pituitary cells at concentrations of 10(-10) and 10(-9) M (P less than 0.025). The specificity of GRF for GH control, whether stimulatory or inhibitory, was seen by the failure of GRF to modify PRL, TSH, or LH release. Our results indicate that injection of larger doses of GRF into the 3V produce GH release, but at lower doses, 3V GRF may exert an action centrally to inhibit GH release. We propose that hypothalamic GRF may decrease its own neurosecretion by negative ultrashort loop feedback.

Evidence for a physiological role for oxytocin in the control of prolactin secretion.

The presence of oxytocin (OT) in neuronal elements of the external layer of the median eminence and in hypophysial portal plasma suggests a role for the peptide in the control of anterior pituitary function. We have reported previously that OT stimulates PRL release in vitro; therefore, we attempted to establish evidence for a physiological PRL-releasing role for OT. Plasma OT levels rose significantly just before the PRL surges occurring during a suckling stimulus in lactating rats (10 min after pup reinstatement vs. 15 min for PRL) and 48 h after estrogen injection in ovariectomized (OVX) rats (at 1200 h vs. 1300 h). Dispersed anterior pituitary cells harvested from lactating female rats and OVX estrogen-primed rats released PRL in a specific, significant, and dose-related fashion when perifused in vitro with incubation medium containing 10(-7)-10(-9) M OT, doses similar to levels found previously in hypophysial portal plasma. Infusion of antiserum specific for OT into lactating females before pup reinstatement and into estrogen-primed OVX rats 2 h before the expected release of endogenous OT delayed and significantly reduced subsequent PRL surges compared to levels in saline-or normal rabbit serum-infused rats; however, PRL release was not completely abolished. These data indicate that OT plays a physiological role in the hypothalamic control of PRL secretion and further suggest the importance of multiple factors in coordinated regulation of PRL release.

Neuropeptide-Y suppresses pulsatile secretion of luteinizing hormone in ovariectomized rats: possible site of action.

The hypothesis that neuropeptide-Y (NPY) suppresses pulsatile LH secretion in ovariectomized (OVX) rats was examined. Rats were bilaterally OVX and 6 weeks (Exp 1) or 2 weeks (Exp 2) later a stainless steel cannula was implanted in the third cerebral ventricle (3V). Seven to 10 days later, an intraatrial cannula was inserted. The next day, a blood sample was withdrawn, and each conscious unrestrained animal received a 3V injection of synthetic porcine NPY (5 or 0.5 micrograms/2 microliters saline) or vehicle in Exp 1. Blood samples were taken every 10 min for 2 h and centrifuged, and the plasma was analyzed for LH by RIA. In Exp 2, OVX rats received a 3V injection of NPY (5 micrograms/2 microliters) or vehicle. Blood samples were taken before and 60 min after injection. At 60 min, LHRH (10 ng/100 g BW) was injected iv, and blood was withdrawn 10, 20, 60, and 120 min later. NPY caused a dramatic dose-related reduction in the pulsatile release of LH compared to that in vehicle-treated rats. The 5.0-micrograms dose of NPY significantly reduced LH pulse frequency (P less than 0.05), pulse amplitude (P less than 0.01), and trough levels (P less than 0.01) compared to those in saline-injected controls. The lower dose of NPY (0.5 micrograms) significantly decreased the mean LH levels throughout the 2-h sampling period and slightly, though not significantly, the pulse frequency. Administration of LHRH increased plasma LH levels by 124% in control animals and by 1239% in NPY-injected rats. The results of these studies indicate that the suppressive effects of NPY on pulsatile LH release appear to be exerted through inhibition of pulsatile LHRH secretion from the hypothalamus.

Hypothalamic and pituitary sites of action of oxytocin to alter prolactin secretion in the rat.

To determine whether oxytocin (OT) could alter the release of PRL and other hormones from the anterior pituitary gland, the effects of OT were examined in two in vitro and two in vivo test systems. Cells dispersed from anterior pituitary glands of intact adult male rats were incubated in medium containing OT at doses of 10(-8), 10(-7), 10(-6), and 10(-5) M in two trials. OT stimulated PRL release 1.5-fold (P less than 0.01) and 2- to 3-fold (P less than 0.001) above control levels at 10(-8) and 10(-7) M doses, respectively, thus indicating a dose-dependent relationship. Higher doses did not produce a further elevation above that obtained with 10(-7) M OT. Arginine vasopressin (AVP) caused a slight decrease in PRL release from dispersed cells while TRH produced a small (25%), significant, but nondose-related increase in PRL release. Hemipituitary glands from adult male rats, incubated with 10(-6) and 10(-5) M OT, released twice as much PRL (P less than 0.01) into the medium as paired controls, but 10(-7) M OT was ineffective. The iv injection of 1 or 10 micrograms OT into conscious male rats elevated plasma PRL by 50% (P less than 0.05) or 500% (P less than 0.001), respectively, above basal values at 5 min only. Vehicle or 0.1 microgram OT were without effect. When 0.1 microgram OT was microinjected into the third ventricle (3V) of conscious male rats, it paradoxically reduced plasma PRL by 40% at 30 min (P less than 0.05), whereas 1 microgram OT significantly lowered PRL at 5-60 min, with the maximum suppression (60%, P less than 0.001) occurring at 30 min. These latter findings may indicate that an ultrashort loop feedback mechanism exists whereby exogenous OT decreases hypothalamic OT secretion, thereby reducing the OT stimulus for PRL release. The specificity of the OT effect on PRL was attested to by the failure of OT to alter significantly FSH, LH, and TSH in each system. GH was unchanged except that 3V-injected OT (1 microgram only) elevated (P less than 0.001) plasma GH at 15-30 min. These results support the view that OT acts directly on the cells of the anterior pituitary gland at low to high doses to release PRL specifically and in a dose-related fashion. In contrast, 3V injection of OT reduces PRL secretion, thereby suggesting that OT may decrease its own neurosecretion by ultrashort loop feedback and thus reduce an OT stimulus for PRL release.

Endotoxin-induced suppression of the somatotropic axis is mediated by interleukin-1 beta and corticotropin-releasing factor in the juvenile rat.

This study extends the neuroendocrine role of central interleukin-1 beta (IL-1 beta) during the stress of lipopolysaccharide (LPS) challenge to include inhibition of the somatotropic [GH-releasing hormone (GHRH)-somatostatin (SRIF)-GH] axis in juvenile male rats and clarifies the role of CRF in the mediation of LPS/IL-1-induced changes in GHRH and SRIF neurosecretion. The results of the in vivo component of this study demonstrated that LPS treatment (2.5 mg/kg twice daily for 5 days) caused a significant attenuation of body weight gain for 2 days (2.4 +/- 1.7% vs. 10.3 +/- 1.8% BW/day in saline controls; P < 0.05) and failure of catch-up growth thereafter even though a small transient suppression of food intake returned to normal by the second of 4 days of treatment. Associated with the first day of growth attenuation was an acute suppression of all plasma GH parameters, including GH mass (area under the curve, 1.972 +/- 0.1837 vs. 6.402 +/- 1.7 micrograms/ml.6 h for saline controls; P < 0.05), in animals receiving an acute bolus of LPS, which was blocked by prior microinjection of IL receptor antagonist protein (IRAP) into the third ventricle. In contrast, GH parameters associated with the second day of LPS-suppressed body weight gain were increased (GH mass, 9.4 +/- 2.2 vs. 3.5 +/- 0.5 micrograms/ml.4 h in saline controls; P < 0.05). These increases were reversed after another 2 days of LPS treatment. In a series of in vitro experiments using medial basal hypothalamic (MBH) explants incubated with LPS [100 ng/ml alone or with 10(-7) M IRAP or 10(-6) M CRF antagonist (CRF-ANT)], GHRH release from MBH incubated with LPS was significantly greater than that in controls (231 +/- 79% vs. 71 +/- 34% of baseline release; P < 0.05), and this stimulation was antagonized by both IRAP and CRF-ANT. SRIF release was significantly increased by incubation with LPS (163 +/- 28% vs. 97 +/- 20% of the baseline for controls; P < 0.05) and blocked (to 88 +/- 14% of the baseline) by IRAP, but not by CRF-ANT. Finally, when MBH explants were incubated with IL-1 beta (10(-9) M), there was a significant inhibition of in vitro GHRH release (37.9 +/- 6.7% vs. 74.9 +/- 16.6% for controls), which was reversed by IRAP and CRF-ANT, and a significant stimulation of SRIF release (168.7 +/- 37.5% vs. 98.0 +/- 11.6% for controls), which was reversed by IRAP, but not CRF-ANT.(ABSTRACT TRUNCATED AT 400 WORDS)

Effect of a growth hormone-releasing factor antagonist on compensatory renal growth, insulin-like growth factor-I (IGF-I), and IGF-I receptor gene expression after unilateral nephrectomy in immature rats.

We have recently reported that pulsatile GH secretion is elevated 24 h after unilateral nephrectomy (UNX) in adult rats. In addition, suppression of the increase in GH with an antagonist to GH-releasing factor (GRF-AN) significantly attenuated compensatory renal growth (CRG) in adult rats. The present study examined the role of GH in CRG in immature animals. Pulsatile GH release was determined 24 h post-UNX in immature (26-28 days of age) sham-operated and UNX male Wistar rats. In contrast to the adult UNX rats, no increase in GH secretion was seen in the immature UNX rats compared with that in the controls. When pulsatile GH release was suppressed with GRF-AN, there was preferential growth of the remnant kidney despite the attenuated gain in whole body weight. In addition, insulin-like growth factor-I (IGF-I) and IGF-I receptor mRNA levels were elevated 3-fold in the remnant kidneys of GRF-AN-treated rats, despite the suppression of pulsatile GH release. These findings suggest that the initial phase of CRG is GH independent in the immature rat and, further, that CRG is associated with an increase in IGF-I and IGF-I receptor gene expression that is independent of episodic GH secretion.

Stimulation of serotonin1A receptors increases release of prolactin in the rat.

The effect of serotonin1A receptor agonists on release of prolactin was examined in awake, freely-moving male rats in which a catheter in the jugular vein allowed samples of blood to be collected periodically after intravenous injection of the agonist. The serotonin1A receptor agonist, 8-hydroxy-2(di-n-propylamino) tetralin (8-OHDPAT) increased concentrations of prolactin in plasma rapidly and in a dose-related manner. Concentrations of prolactin peaked within 9 min after intravenous injection of 8-OHDPAT and returned to baseline values within 30 min. Another serotonin1A receptor agonist, 5-methylurapidil (5-MeU), produced a similar response of prolactin. The effects of these agonists on release of prolactin were completely blocked by pretreatment with the serotonin receptor antagonists, methysergide and metergoline, administered 1 or 2 hr before the agonist. These results demonstrated that serotonin1A receptors can mediate the effects of serotonin on release of prolactin in the male rat.

Prenatal exposure to benzodiazepine--I. Prenatal exposure to lorazepam in mice alters open-field activity and GABAA receptor function.

Prenatal exposure to benzodiazepines may lead to developmental abnormalities in humans and animals. To assess the behavioral and neurochemical effects of such exposure, pregnant mice were treated with lorazepam, 2 mg/kg/day, from days 13-20 of gestation, and open-field activity was assessed in offspring at 3 and 6 weeks of age and the function of GABAA receptors at 6 weeks of age. Activity was increased in mice exposed to lorazepam, compared to untreated or vehicle-treated controls at 3 weeks, but was unchanged at 6 weeks. Muscimol-stimulated uptake of chloride was decreased in lorazepam-treated mice, compared to controls, with a decrease in maximum uptake but no change in the EC50 for muscimol. Concentrations of lorazepam in maternal plasma and brain showed a similar brain:plasma ratio as previously reported and concentrations in fetal brain were about 50% of maternal levels. Lorazepam persisted for 48 hours after birth in dams but not in the offspring. These results indicate persistent behavioral and neurochemical alterations after prenatal exposure to lorazepam. This model may be useful in assessing other effects of prenatal exposure to benzodiazepine.

Augmentation of GABAA receptor function by chronic exposure to GABA-neutral and GABA-negative benzodiazepine ligands in cultured cortical neurons.

Chronic benzodiazepine agonist administration may lead to decreases in gamma-aminobutyric acidA (GABAA) receptor binding and function, but little information is available concerning chronic GABA-neutral or GABA-negative benzodiazepine exposure. We evaluated effects of chronic exposure to flumazenil (Ro15-1788) and FG 7142 (N-methyl-beta-carboline-3-carboxamide) on GABA-dependent chloride uptake in chick cerebral cortical neurons in primary culture. Acute flumazenil treatment (1 microM) had no effect on chloride uptake, but uptake was increased after 2 days of exposure. Similar increases were observed after 4 and 10 days. Flumazenil, 0.1 microM, had no effect after 10 days, and a 10 microM concentration had a similar effect as the 1 microM concentration. Acute FG 7142 (1 microM) decreased chloride uptake, but uptake was increased markedly after 2, 4, and 10 days of treatment. No effect was observed after treatment for 10 days with 0.1 microM, but a 10 microM concentration showed similar enhancement to the 1 microM concentration. Concurrent treatment with 0.3 microM flumazenil which did not affect chloride uptake and 1 microM FG 7142 for 10 days substantially attenuated the effects of FG 7142, suggesting that FG 7142 effects are mediated at the benzodiazepine site. Benzodiazepine receptor binding was increased in cultures treated for 10 days with 1 microM flumazenil or FG 7142, with an increase in receptor number in both cases but no change in apparent affinity. Neither flumazenil nor FG 7142 (1 microM for 10 days) altered GABA-independent chloride uptake, total cellular protein, protein synthesis or degradation, or neuronal survival. These results indicate that both chronic GABA-neutral and GABA-negative benzodiazepine exposures in cultured cortical neurons lead to increases in GABA-dependent chloride uptake and benzodiazepine binding. Effects of GABA-negative benzodiazepine exposure appear to be greater than those observed with GABA-neutral benzodiazepine exposure.

Differential hypothalamic control of FSH secretion: a review.

There are many circumstances in which the release of FSH and LH is dissociated; however, many of these are now thought to be brought about by interactions of LH-releasing hormone (LHRH), which stimulates not only LH but also FSH release, and the gonadal peptide, inhibin, which acts at the pituitary to suppress FSH release selectively. There are also many examples which can only be explained by postulating separate hypothalamic control of FSH and LH release. For example, electrochemical stimulation of the medial preoptic area elicited only LH release, whereas stimulation further caudally elicited equivalent LH release but FSH release as well. Points of stimulation particularly in the dorsal anterior hypothalamic area (DAHA) evoked only FSH release. Furthermore, implantation of prostaglandin E2 in various hypothalamic loci in a region extending from the DAHA caudally and ventrally to the caudal median eminence (ME) selectively elicited FSH release. Lesions of the DAHA resulted in a decrease of plasma FSH but not LH in castrated male and female rats and also suppressed the post-castration rise in FSH in males. In ovariectomized estrogen-primed rats with DAHA lesions, injection of progesterone provoked a normal LH surge but a significantly depressed FSH surge. Anterior ME lesions in castrates lowered LH levels more than FSH levels. Extracts of the DAHA evoked greater FSH and LH release in vitro than could be accounted for by the content of LHRH in the extracts, but there was no preferential release of FSH. On the other hand, extracts of the organum vasculosum lamina terminalis (OVLT) evoked dramatically increased FSH release above that which could be accounted for by the content of LHRH. Lastly, posterior ME extracts had more FSH-releasing activity than could be accounted for by their content of LHRH. All these results suggest the existence of an FSH-releasing factor (FSHRF) and lead to the speculation that the cell bodies of FSHRF neurons are located in the DAHA, with axons projecting to the OVLT and to the posterior ME. In other experiments, attempts were made to purify rat and sheep hypothalamic extracts by gel filtration on Sephadex G-25 and to assay the FSH-releasing activity by both bio- and immunoassay. Using this approach, we obtained evidence for the early emergence of a bioactive FSHRF prior to the emergence of LHRH from the column. Although much more work remains to be done, the accumulated evidence strongly supports the concept of a distinct FSHRF.