Robert V. Gallo

Stimulatory or Inhibitory Effects of Angiotensin II upon LH Secretion in Ovariectomized Rats: a Function of Gonadal Steroids

The effects of intraventricular infusions of artificial cerebrospinal fluid (aCSF) or angiotensin II (AII) on LH secretion were investigated in rats that had been ovariectomized for 8 days. In untreated ovariectomized rats, the mean whole blood concentration of LH as well as the amplitude, frequency, and nadir of the LH pulses were not affected by infusion of aCSF or 15 ng AII/h, but were suppressed in a dose-dependent fashion by infusion of AII at doses of 150 or 600 ng/h. The AII receptor antagonist, saralasin, blocked the inhibitory effect of AII, demonstrating the specificity of the response to AII. In ovariectomized rats pretreated with estradiol, infusion of AII did not modify mean blood LH levels. However, in ovariectomized rats pretreated with both estradiol and progesterone, infusions of AII at 150 or 600 ng/h produced dose-dependent increases in mean LH concentrations. The results demonstrate both inhibitory and stimulatory effects of AII upon LH secretion, the direction of the effect being determined by gonadal steroids.

Medial preoptic area involvement in norepinephrine-induced suppression of pulsatile luteinizing hormone release in ovariectomized rats.

This study examined whether the medial preoptic area (MPOA) is a site which mediates the inhibitory effects of norepinephrine (NE) on pulsatile luteinizing hormone (LH) secretion in ovariectomized rats. Animals were bled continuously at a rate of 50 microliter whole blood/7 min for 2 h prior to push-pull perfusion in the MPOA, and during a 2-3 h period of perfusion of the MPOA (20 microliter/min) with artificial CSF, or 2 or 20 pg NE/min. In another group of rats LH levels were only determined during a 2-3 h period of MPOA perfusion with CSF. Pulsatile LH release was not affected by push-pull perfusion with CSF when a comparison was made to preperfusion LH values in the same rats. Moreover, LH levels obtained from rats only bled during MPOA perfusion with CSF were not different from LH values obtained during the preperfusion periods in the other groups. However, push-pull perfusion of the MPOA with 2 or 20 pg NE/min significantly suppressed mean LH levels by causing a 35-45% reduction in pulse frequency. No decrease occurred in LH pulse amplitude. Therefore, these studies demonstrate that NE acting at the level of the MPOA can suppress pulsatile LH release solely by decreasing LH pulse frequency.

Kappa-Opioid Receptor Involvement in the Regulation of Pulsatile Luteinizing Hormone Release During Early Pregnancy in the Rat

The object of this study was to gain further insight into endogenous opioid peptide suppression of pulsatile luteinizing hormone (LH) release in early gestation in the rat by examining whether selective blockade of mu ‐, delta ‐, or kappa‐opioid receptor(s) results in stimulation of pulsatile LH secretion at this time. Previous reports demonstrated stimulation of pulsatile LH release during early gestation by intravenous infusions of naloxone, an endogenous opioid peptide receptor antagonist whose binding is not specific to a single class of opioid peptide receptors. In the present study, naloxone infused intraventricularly similarly stimulated an increase in pulsatile LH release on Days 7 to 8 of gestation. Antagonists of specific opioid peptide receptor subtypes were thus given by this route. Administration of nor‐binaltorphimine, an antagonist of kappa‐opioid receptors, but not β‐funaltrexamine or ICI 174, 864, antagonists of mu‐ and delta‐opioid receptors, respectively, exerted a stimulatory action on both LH pulse amplitude and frequency similar to that of naloxone, indicating involvement of this opioid peptide receptor subtype in the endogenous opioid peptide suppression of pulsatile LH release in early gestation in the rat.

Further Studies on Norepinephrine-Induced Suppression of Pulsatile Luteinizing Hormone Release in Ovariectomized Rats

The present study examined three aspects of the inhibitory effects of continuous intraventricular infusion of norepinephrine (NE) on pulsatile luteinizing hormone (LH) release in ovariectomized, nonsteroid-primed rats: whether the inhibitory effects of NE infusion were exerted on LH pulse frequency and/or amplitude; whether central nervous system desensitization occurred in response to the inhibitory effects of continuous NE infusion on pulsatile LH secretion, and whether dopamine of serotonin were involved as possible interneuronal transmitter mediators of NE-induced suppression of pulsatile LH release. Unanesthetized rats with external jugular cannulae were bled continuously at a rate of 50 microliters whole blood/7 min for 2 h prior to infusion and for 2-3 h during continuous intraventricular infusion of artificial cerebrospinal fluid or NE. Infusion of cerebrospinal fluid had no effect on pulsatile LH release, while continuous infusion of 0.3 or 1.8 micrograms NE/h for 2-3 h produced suppression of pulsatile LH secretion. Although desensitization to the stimulatory effects of NE on LH release in ovariectomized, steroid-primed rats had been observed to occur rapidly within 90 min after the onset of infusion, desensitization to the inhibitory effect of NE on pulsatile LH release did not occur even after continuous infusion of NE for periods up to 20 h. Mean blood LH levels were as low in rats bled 17-20 h after the onset of NE infusion as in those bled at 0-3 h. The suppressive effect of NE on pulsatile LH release was not prevented by prior blockade of dopamine or serotonin receptors with pimozide or metergoline, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Endogenous Opioid Peptide Regulation of Pulsatile Luteinizing Hormone Secretion during Pregnancy in the Rat

The objective of this study was to determine if endogenous opioid peptides (EOPs) influence the pattern of pulsatile luteinizing hormone (LH) secretion on days 6-8, 14-16 and 22 of gestation in the rat. Unanesthetized animals with two jugular cannulae were initially infused with 0.9% saline during which the control pattern of pulsatile LH release was determined. Possible EOP involvement was then determined by infusion of the EOP receptor antagonist naloxone. Plasma estradiol (E2) and progesterone (P) values increased between days 6-8 and 14-16. While plasma E2 values remained elevated through day 22, plasma P values declined by 90%. As previously reported, mean blood LH levels during the control period on day 22 were higher than on days 6-8 and 14-16 due to an increase in LH pulse frequency. At each stage of gestation naloxone infusion increased mean blood LH levels. This stimulatory action of naloxone was reduced in a dose-dependent fashion by simultaneous infusion with morphine, demonstrating that this effect is mediated via EOP receptors. There was no difference in the in vivo pituitary responsiveness to LH-releasing hormone (LHRH) between rats infused with saline or naloxone at any stage of pregnancy, demonstrating that the stimulatory effect of naloxone was not exerted at the pituitary level. Naloxone increased both the amplitude and frequency of pulsatile LH secretion on days 6-8, and stimulated frequency on days 14-16. The effect on amplitude could not be assessed on days 14-16 because too few rats exhibited pulsatile LH secretion prior to naloxone infusion. The increase in pulse frequency was similar on days 6-8 and 14-16. Although naloxone increased LH pulse amplitude and frequency on day 22, these increases were significantly less than those seen on days 6-8 and 14-16, respectively. Pituitary responsiveness to LHRH was less at all stages of pregnancy in comparison to responsiveness in ovariectomized rats, and progressively declined from days 6-8 through day 22. The lowest responsiveness to LHRH was seen on day 22 and contributed, at least in part, to the diminished increase in LH pulse amplitude in response to naloxone infusion on day 22 compared to days 6-8. The reduced naloxone-induced increment in LH pulse frequency on day 22, occurring coincident with a precipitous decline in plasma P levels, suggests a decreased EOP suppression of pulse frequency at this time.(ABSTRACT TRUNCATED AT 400 WORDS)

Medial preoptic-anterior hypothalamic area involvement in the suppression of pulsatile LH release by a mu-opioid agonist in the ovariectomized rat

The objective of this study was to examine whether specific activation of mu-opioid receptors at the level of the medial preoptic-anterior hypothalamic area (MPOA-AHA) could suppress pulsatile LH release. The experiments were done using rats that had been ovariectomized (OVX) 24 hr before on diestrus 2, animals in which we have previously demonstrated an active endogenous opioid peptide suppression of pulsatile LH release (2). DAGO, DPDPE, or U50488H, specific agonists of mu-, delta- and kappa-opioid receptors, respectively, were continuously applied directly to the MPOA-AHA by means of push-pull perfusion. Perfusion of the MPOA-AHA with 0.5 micrograms DAGO/hr suppressed LH pulse amplitude. This effect of DAGO was not due to spread to the third ventricle and subsequent diffusion via the CSF to another CNS site, since push-pull perfusion with this dose of DAGO in the region just dorsal to or in the posterior hypothalamus was ineffective in altering LH pulse amplitude. The response to DAGO was dose-dependent since a higher dose (4.8 micrograms/hr) markedly suppressed both LH pulse amplitude and frequency. The same doses of DPDPE and U50488H (0.5 and 4.8 micrograms/hr) had no effect on pulsatile LH secretion, providing support for mu receptor involvement in the DAGO-induced suppressive action. These data demonstrate MPOA-AHA involvement in the suppression of pulsatile LH release by a mu-opioid agonist in the OVX rat.

Steroid-independent endogenous opioid peptide suppression of pulsatile luteinizing hormone release between estrus and diestrus 1 in the rat estrous cycle

We have previously demonstrated an absence of ovarian steroid negative feedback on pulsatile luteinizing hormone (LH) release between estrus and early diestrus 1 (D1) in the rat estrous cycle. The object of the present study was to determine if there was a steroid-independent endogenous opioid peptide (EOP) suppression of pulsatile LH release in this same 24-h interval, and if so, which parameter(s) of pulsatile LH release were affected. Rats were bled on estrus, or 24 h following sham ovariectomy (OVX), or OVX at 08.30-10.00 h on estrus. At the time of bleeding all rats were infused i.v. for 4 h either with 0.9% saline (0.5 ml/h) or naloxone (0.005, 0.05, 0.5, or 2 mg/kg/h). At 1 h after the infusion began, rats were bled for 3 h (40 or 50 microliters whole blood/5 min) between 09.30 and 12.30 h. Mean blood LH levels increased between estrus and early D1 due to increases in LH pulse amplitude and frequency. OVX on estrus decreased plasma levels of estradiol and progesterone 24 h later, but did not augment the increase in pulsatile LH release. However, naloxone infusion augmented the increase in pulsatile LH secretion in sham ovariectomized rats in a dose-dependent fashion. While infusion of 0.005 or 0.05 mg/kg/h had no effect, 0.5 or 2 mg/kg/h increased blood LH levels by increasing both LH pulse amplitude and frequency. The stimulatory effect of naloxone on pulsatile LH release was blocked by simultaneous infusion of morphine, demonstrating that the effect was mediated by EOP receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of prodynorphin-derived opioid peptides on the ovulatory luteinizing hormone surge in the proestrous rat.

The objective of this study was to determine whether prodynorphin-derived opioid peptides could block the spontaneous luteinizing hormone (LH) surge and ovulation, and if so, whether this inhibitory action was mediated through κ-opioid receptors. Various doses of dynorphin peptides (dynorphin A1–17, dynorphin A1–8, dynorphin B, α- and β-neoendorphin) were infused into the brain through third-ventricle cannulae in rats between 1330–1800 h on proestrus. Each dynorphin peptide blocked the LH surge and ovulation in a dose-dependent manner. Dynorphin A1–17 and A1–8 were equally effective in producing these actions, and more potent than either dynorphin B or α- or β-neoendorphin. U50,488H, a specific κ-opioid receptor agonist, also blocked the LH surge and ovulation. When a mixture of five dynorphin peptides was infused intraventricularly, each at a dose that inhibited the LH surge, both the surge and ovulation were blocked. However, when norbinaltorphimine, a specific κ-opioid receptor antagonist, was coinfused with the mixture of dynorphin peptides, the LH surge and ovulation were fully restored. These results demonstrate that prodynorphin-derived opioid peptides, acting through κ-opioid receptors, can block the LH surge and ovulation. Dynorphin A1–17 and A1–8 are the most potent in this regard.

Neurotransmitter involvement in naloxone-induced stimulation of pulsatile LH release on day 8 of pregnancy in the rat

Naloxone, an endogenous opioid peptide (EOP) receptor antagonist, increases LH pulse frequency and amplitude in early gestation in the rat (7). The object of this study was to further explore the suppression of pulsatile LH release by EOPs on day 8 of pregnancy by examining whether inhibition of norepinephrine (NE) or epinephrine (EPIN) synthesis, or stimulation of gamma-aminobutyric acid (GABA)-B receptors, modified the ability of naloxone infusion (0.5 mg/kg/hr for 3.5 hr) to stimulate pulsatile LH secretion. Blood sampling (50 microliters whole blood/5 min) began 0.5 hr after the onset of infusion. Three studies were conducted. 1) LY 134046 (PNMT inhibitor, 50 mg/kg IP), given at -27, -20, and -3 hr relative to the onset of a 3-hr blood sampling period, produced no change in hypothalamic-preoptic area (HPOA) levels of NE, and a 76% decline in HPOA-EPIN levels, as determined by HPLC. Although basal LH pulse frequency was reduced, this treatment had no effect on the stimulatory action of naloxone on pulsatile LH release. 2) FLA-63 (DBH inhibitor, 25 mg/kg IP) given at -3 hr produced a 72% decline in HPOA-NE levels, a 44% decrease in HPOA-EPIN values, and blocked the stimulatory action of naloxone on LH pulse frequency. Since depletion of HPOA-EPIN by LY 134046 did not compromise the LH response to naloxone, this effect of FLA-63 is due to depletion of HPOA-NE levels. 3) While saline had no effect on the increased pulsatile LH release caused by naloxone, administration of baclofen (GABA-B receptor agonist, 6 mg/kg IV) suppressed the pulsatile LH secretory response to naloxone infusion.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of estradiol and progesterone on pulsatile LH secretion in 8-day ovariectomized rats

Previous studies in our laboratory [Endocrinology 114: 1605-1612 (1984); Neuroendocrinology 41: 252-257 (1985)] examined the influence of ovarian steroids on pulsatile luteinizing hormone (LH) release, and involved immediate replacement following ovariectomy (OVX) of estradiol (E2) and progesterone (P) for a 24-hour period within the physiological context of the estrous cycle. The present study investigated the effects of replacing E2 and/or P 1 week after OVX, and therefore examined whether the time elapsed following OVX influences the effects of ovarian steroids on pulsatile LH release. Immediately after jugular venous cannulation, rats were implanted with either empty silastic capsules or capsules capable of restoring physiological levels of E2 and/or P comparable to those found in intact rats between the intervals of diestrus 1 (D1) and diestrus 2 (D2), or D2 and proestrous morning. 24 h later, these rats were bled continuously at a rate of 50 microliters whole blood/6 min for 3 h for analysis of pulsatile LH secretion. Rats with empty capsules had decreased levels of E2 and P and elevated mean blood LH levels, pulse amplitudes and frequencies. Two groups of animals with E2 capsules had plasma E2 levels comparable to those seen either in the D1-D2 or D2-proestrous intervals, decreased levels of P, and in both cases significant decreases in LH pulse amplitude, but no change in LH pulse frequency or basal LH secretion. Since mean blood LH levels in 8-day ovariectomized rats are determined by LH pulse amplitude, frequency and basal LH secretion [Neuroendocrinology 37: 421-426 (1983)].(ABSTRACT TRUNCATED AT 250 WORDS)