Acute effects of bromocriptine, cyproheptadine, and valproic acid on plasma adrenocorticotropin secretion in Nelson's syndrome.

Previous studies have found that bromocriptine, cyproheptadine, and valproic acid can reduce ACTH secretion in Nelson's syndrome, but none of these agents has achieved widespread use due to their failure to normalize ACTH in most patients. The current study was undertaken to determine whether these three agents, which act through different mechanisms, decrease plasma ACTH synergistically when administered together. Six adult female patients (mean age, 41 yr) with Nelson's syndrome were studied. ACTH was measured every 20 min for 8 h, 2 h before and 6 h after each of the following six treatments: placebo, bromocriptine (2.5 mg), cyproheptadine (8 mg), valproic acid (1 g), cyproheptadine plus valproic acid, and the combination of all three drugs. The sequence of treatments was determined randomly, and there was an interval of at least 2 days between each treatment. The hourly ACTH values were averaged, and the percent maximal suppression of plasma ACTH, relative to the baseline values before drug administration, was compared among the six treatments. Basal plasma ACTH levels in the six patients ranged from 40-3324 pmol/L (normal range, 1-8). The percent maximal suppression of ACTH after administration of placebo (6 +/- 11%), cyproheptadine (17 +/- 15%), valproic acid (37 +/- 10%) or the combination of cyproheptadine and valproic acid (19 +/- 14%) did not achieve statistical significance. Bromocriptine, on the other hand, caused a significant decrease in plasma ACTH (52 +/- 8%; P < 0.05), as did the combination of bromocriptine, cyproheptadine, and valproic acid (58 +/- 12%; P < 0.05). However, the combined effect of the three drugs did not significantly exceed the effect of bromocriptine alone. We conclude that at the doses studied, bromocriptine had the greatest acute effect in suppressing ACTH secretion in Nelson's syndrome, and that combined administration with valproic acid and cyproheptadine did not further increase this acute ACTH-suppressive effect.

Localization of vasopressin mRNA and immunoreactivity in pituicytes of pituitary stalk-transected rats after osmotic stimulation.

The presence of [arginine] vasopressin (AVP) mRNA and AVP immunoreactivity in pituicytes of the neural lobe (NL) of intact and pituitary stalk-transected rats, with and without osmotic stimulation, was examined. AVP mRNA was analyzed by Northern blotting, as well as by in situ hybridization in combination with immunocytochemistry using anti-glial fibrillary acidic protein (GFAP) as a marker for pituicytes. In intact rats, a poly(A) tail-truncated 0.62-kb AVP mRNA was detected in the NL and was found to increase 10-fold with 7 days of continuous salt loading. Morphological analysis of the NL of 7-day salt-loaded rats revealed the presence of AVP mRNA in a significant number of GFAP-positive pituicytes in the NL and in areas most probably containing nerve fibers. Eight days after pituitary stalk transection the NL AVP mRNA diminished in animals given water to drink, whereas in those given 2% saline for 18 h followed by 6 h of water, a treatment repeated on 6 successive days beginning 2 days after surgery, the 0.62-kb AVP mRNA was present. The AVP mRNA in the pituitary stalk-transected, salt-loaded rats showed an exclusive cellular distribution in the NL, indicative of localization in pituicytes. Immunoelectron microscopy showed the presence of AVP immunoreactivity in a subpopulation of pituicytes 7 and 10 days after pituitary stalk transection in salt-loaded animals, when almost all AVP fibers had disappeared from the NL. These data show that a subset of pituicytes in the NL is activated to synthesize AVP mRNA and AVP in response to osmotic stimulation.

The effect of salt-loading on corticotropin releasing hormone and arginine vasopressin mRNA levels in the mouse hypothalamus: a quantitative in situ hybridization analysis.

Previously, we showed that during salt-loading in mice there was an acute rise in plasma ACTH levels after 2 days followed by a transient decrease after 4 and 9 days. Pro-opiomelanocortin (POMC) mRNA levels in the anterior pituitary increased after 2 days and returned to normal thereafter. In this study, changes in hypothalamic CRH and AVP mRNA levels during salt-loading were investigated using quantitative in situ hybridization histochemistry. CRH mRNA was expressed only in the paraventricular nucleus (PVN), while AVP mRNA was expressed in both the supraoptic (SON) and paraventricular nuclei. CRH mRNA levels were unchanged after 2 days salt-loading, but declined to 77% of control levels after 9 days. AVP mRNA levels rose to 260% and 634% of control levels in the SON, and to 352% and 522% of control levels in the PVN, after 2 and 9 days salt-loading, respectively. These data suggest a major role of AVP in the acute stimulation of ACTH secretion and POMC mRNA levels seen after 2 days salt-loading. Desensitization of AVP receptors at the corticotroph level and a centrally mediated inhibition of CRH release may account for the decrease of ACTH secretion and POMC mRNA levels in the anterior pituitary with prolonged salt-loading.

Rat mitochondrial coupling factor 6: molecular cloning of a cDNA encoding the imported precursor.

A cDNA clone encoding the precursor to the rat mitochondrial protein coupling factor 6 (F6) has been isolated and sequenced. The deduced amino acid sequence of the rat precursor protein shows 78% and 74% identity with the human and bovine F6 pre-proteins, respectively.

Coordinate regulation of mRNA levels of pro-opiomelanocortin and the candidate processing enzymes PC2 and PC3, but not furin, in rat pituitary intermediate lobe.

Pro-opiomelanocortin is a multivalent hormone precursor which is processed at pairs of basic residues in a tissue-specific manner to release biologically active peptides. We have examined the message levels of three candidate pro-opiomelanocortin processing enzymes in the intermediate lobe of the rat pituitary following treatment with a dopamine receptor agonist and antagonist which are known to regulate pro-opiomelanocortin mRNA levels. Message levels for PC2 and PC3 but not furin were coordinately regulated with pro-opiomelanocortin transcripts supporting a role for PC2 and PC3 in the maturation of the pro-opiomelanocortin precursor in the rat pituitary intermediate lobe.

Changes in proopiomelanocortin messenger ribonucleic acid levels in the rostral periarcuate region of the female rat during the estrous cycle.

GnRH synthesis and release are regulated by a number of neurotransmitter systems. Several studies have implicated the opioidergic system as one of the important modulators of GnRH. To obtain an index of the activity of beta-endorphin-secreting neurons during the estrous cycle, we measured levels of proopiomelanocortin mRNA (POMC mRNA) in the periarcuate region at different cycle stages, using in situ hybridization. Ten female Sprague-Dawley rats (200-230 g) were killed at each of 11 times during the 4-day estrous cycle. Fresh frozen sections were made through the rostral arcuate nucleus and placed on gelatin-coated slides. A 48-base probe complementary to rat POMC mRNA was 3' end-labeled with [35S]dATP and applied to individual sections in hybridization buffer. Sections were washed and exposed to film. Relative amounts of POMC mRNA were measured by obtaining optical densities with an image analyzer. POMC mRNA levels varied significantly. At proestrus, they were low just before the onset of the LH surge, followed by a sharp rise that afternoon. On the day of estrus, POMC mRNA remained elevated and then declined again on metestrus. A second but smaller rise was seen in the late afternoon of metestrus. This pattern of changes in POMC mRNA is consistent with an inhibitory effect of beta-endorphin on GnRH after the midcycle surge and in the postovulatory phase of the cycle, while low levels of POMC mRNA in the early afternoon of proestrus may permit the release of GnRH, which triggers the LH surge. The changes in POMC mRNA approximately parallel changes in progesterone in the cycle.

[3H]2-Nitroimipramine: a selective "slowly-dissociating" probe of the imipramine binding site ("serotonin transporter") in platelets and brain.

Previous studies have demonstrated a close functional and structural relationship between the "high affinity" binding site for [3H]imipramine and the presynaptic and platelet uptake site(s) for serotonin. Recently we have synthesized several nitro derivatives of imipramine which have a very high affinity for the imipramine binding site and which dissociate very slowly when incubations are performed at 0-4 degrees C. In this report, we describe the characteristics of [3H]2-nitroimipramine binding to platelet and brain membranes. Our results support the relative utility of this ligand for studying the impramine binding site (serotonin transporter) since this analogue has both a higher affinity and specific activity than [3H]imipramine. [3H]2-Nitroimipramine by virtue of its extremely slow dissociation rate should be a valuable tool in subsequent characterization and purification of the serotonin uptake or transport site.