α-MSH Concentrations Research Articles

α-MSH and coat color changes in the mouse

The effect of α-MSH on coat color was examined in viable yellow mice (C3H/He-A∗ vy). These mice normally grow a coat of darkly pigmented hair at puberty. This darkening effect was also evident in hair that grew in a region that had been plucked at 13 days of age. Administration of α-MSH increased the darkness of this hair and the hair which grew naturally in an unplucked area. However, the natural coat darkening that occurred at puberty was not associated with an increase in plasma immunoreactive α-MSH levels. Moreover, although bromocryptine, a dopamine agonist that inhibits α-MSH release from the pituitary reduced the darkness of the coat that grew after plucking the reduction in coat darkening was unrelated to changes in plasma α-MSH. Nevertheless, this effect of bromocryptine was reversed when α-MSH was administered together with the drug. Apomorphine had no effect on coat darkening and produced only a slight decrease in plasma α-MSH. Melatonin reduced coat darkening slightly but, like apomorphine, had little effect on plasma α-MSH concentrations. Although α-MSH may have a physiological role in coat darkening in the C3H/He-A∗ vy mouse at puberty the response seems to be unrelated to an increase in circulating α-MSH. Thus, other factors, such as changes in melanocyte sensitivity to α-MSH or inhibitory mechanisms that prevent coat darkening during prepubertal and adult life may be involved in regulation of coat color in the viable yellow mouse.

Regional heterogeneity in the ratio of α-MSH:β-endorphin in rat brain

The molar ratio of α-MSH:β-endorphin varies markedly among discrete microdissected regions of rat brain ranging from 0.57 in the median eminence to 2.74 in the lateral septum. This finding demonstrates that α-MSH and β-endorphin (β-END) are not uniformly distributed in a 1:1 molar ratio in rat brain as one might predict based on the consideration that the two peptides are synthesized in equimolar amounts as part of a common precursor molecule, pro-opiomelanocortin. The data indicate instead that the concentrations of α-MSH and β-END, the two predominant peptides expressed by opiomelantropinergic neurons, are independently regulated in rat brain. The heterogeneity of α-MSH:β-END ratios suggests that the regulation of α-MSH and β-END is regionally specific and may impart functional selectivity to the multisecretory opiomelanotropinergic neuronal system.

Multiple factors involved in the control of ACTH and α-MSH secretion

Alpha 1-adrenergic agents are potent stimulators of ACTH secretion directly at the pituitary level as observed using anterior pituitary cells in primary culture and by in vivo studies. On the other hand, β-adrenergic as well as antidopaminergic agents are also potent stimulators of ACTH secretion but at a suprapituitary level, since no effect of these compounds are observed in vitro. CRF administration has been found to lead to rapid and parallel increases in ACTH and α-MSH concentrations in rat plasma and the mechanism of action of CRF has been studied in vitro using rat anterior and intermediate lobe of pituitary gland for ACTH and α-MSH secretion, respectively. In both cases, CRF stimulates adenylate cyclase activity at ED 50 values of 70 and 350 nM for ACTH and α-MSH, respectively. CRF stimulates adenylate cyelase activity at least partly through a guanyl nucleotide-dependent mechanism. Using rat pars intermedia cells in culture, we have demonstrated the presence, in addition of a CRF receptor, of a β 2-adrenergic receptor which stimulates cyclic AMP accumulation and α-MSH secretion and of a dopaminergic receptor which inhibits cellular activity. These results have been confirmed in in vivo studies where isoproterenol and thioproperazine (a dopamine antagonist) lead to a rapid increase of α-MSH secretion.

α-MSH and zona glomerulosa function in the rat

The responses of rat adrenal zona glomerulosa cells to stimulation by α-MSH and ACTH and related peptides have been studied. The major findings were that: (1) α-MSH stimulated corticosterone production in glomerulosa cells from normal animals at concentrations of about 10 −10 mol/l, but other steroids, including aldosterone, were not significantly stimulated until levels of 10 −7 mol/1 were used. Peptide structure-function relationships showed that in the adrenal cortex, in contrast with other systems, ACTH 4–10 had no effect and did not block the response of glomerulosa cells to α-MSH, bisacetyl Ser 1-α-MSH, (nor-valine-12)-α-MSH, and ACTH 1–13 amide were equipotent with α-MSH, while α-MSH 1–10 had activity but was considerably less potent. α-MSH 6–13, 7–13, 8–13 and lys-11-acetyl-α-MSH were all inactive. N-formyl- N- ϵ-benzyloxycarbonyl α-MSH stimulated only at 10 −6 mol/1. 1. (2) Normalised α-MSH dose-response curves for aldosterone production in glomerulosa cells from normal rats, and corticosterone in inner zone cells were coincident. In glomerulosa cells, prior sodium depletion shifts the dose-response curve for aldosterone to the left, indicating a more sensitive response, and for corticosterone to the right. Bromocriptine treatment (which depresses the level of α-MSH in circulating plasma) and metoclopramide (which enhances it) respectively increased and decreased the sensitivity of the response of corticosterone to α-MSH in subsequently incubated glomerulosa cells, but had no effect on aldosterone. 2. (3) In contrast, normalised ACTH stimulated dose-response curves for glomerulosa corticosterone and aldosterone, and for fasciculata corticosterone production were all coincident, and were unaffected by sodium depletion, or by metoclopramide or bromocriptine pretreatment. 3. (4) Cyclic-AMP production by glomerulosa cells was stimulated by α-MSH only at levels of in excess of 10 −5 mol/1, five orders of magnitude greater than required to produce significant corticosterone stimulation. Under cyclic-AMP stimulation, the normalised responses of glomerulosa corticosterone and aldosterone, and of inner zone corticosterone were all coincident. The data suggest that α-MSH at low concentrations (< 10 −7 mol/1) interacts with a glomerulosa cell receptor which is distinct from the ACTH receptor but interacts with the ACTH receptor at concentrations greater than 10 −5 mol/1. Corticosterone production is stimulated by α-MSH in cells from normal animals at concentrations within the normal range for circulating plasma (~3 × 10 −10 mol/l), while aldosterone is stimulated by similar concentrations of α-MSH in cells from sodium depleted animals. The effects of sodium depletion are not modulated through changes in plasma α-MSH levels. At low concentrations α-MSH stimulation of glomerulosa cells is unlikely to be modulated by cyclic-AMP as second messenger.

α-MSH injected into the septal region reduces fever in rabbits

In previous research the concentration of α-MSH within the septal region of rabbits increased with fever. This finding raises the possibility that the septal concentration of this peptide, which reduces fever when given both peripherally and intracerebroventricularly, is important to limitation of fever. To test this idea, rabbits with cannulas in the septal region were made febrile by IV injections of leukocytic pyrogen (LP). Injection of α-MSH (1 μg bilaterally) into the septal region did reduce fever, consistent with the idea that the increase in septal α-MSH concentration which occurs naturally in fever limits the febrile response. We also noted late rises in body temperature when experimental and control septal injections were given close together in time. These increases in temperature were similar to those known to occur after injections into the primary temperature control in the PO/AH region. This commonality further strengthens the possibility that septal neurons are important to central modulation of body temperature.

Absorption of α-MSH from subcutaneous and intraperitoneal sites in the rat

Pentobarbitone anesthetized rats were injected with 30 nmol (50 μg) α-MSH administered intraperitoneally (IP) and subcutaneously (SC) in an acid-saline vehicle, or SC in a zinc phosphate vehicle. Concentrations of α-MSH in plasma were measured by radioimmunoassay. The pharmacokinetic parameters for the three modes of administration were determined by fitting a one-compartment open model to the plasma level data. The t 1 2 for absorption using the saline vehicle was 7.3 and 5.6 min from the IP and SC sites, respectively. The t 1 2 for absorption from the zinc phosphate complex of 17.7 min was significantly longer. Five percent of the IP dose was absorbed into the systemic circulation giving a peak plasma level of 14.1 nmol/l. The absorption of 2–3 percent was significantly lower following SC administration; peak plasma levels were 8.3 and 4.8 nmol/l for the saline and zinc phosphate vehicles, respectively. The low percentage absorption values indicated a high degree of metabolism of the peptide by peripheral tissues on its passage from the injection sites into the circulation.

Biological and immunological characterization of α-melanocyte-stimulating hormone (α-MSH) in two neuronal systems of the rat brain

Recent immunocytochemical studies have demonstrated the existence of two different neuronal systems containing α-MSH-like material in the brain: one originating from the arcuate nucleus and the other one from the dorsolateral hypothalamus. The aim of the present study was to further characterize α-MSH in these two discrete regions of the rat diencephalon. Intracerebroventricular administration of colchicine resulted in a marked decrease in the number of ACTH and β-endorphin nerve fibers and a significant reduction in ACTH and β-endorphin content in the dorsolateral hypothalamus. Conversely, colchicine treatment did not alter α-MSH, ACTH or β-endorphin content in the arcuate nucleus and did not significantly affect α-MSH concentration in the dorsal region. Reverse-phase high-performance liquid chromatography showed that the major α-MSH-like compound localized in the dorsal hypothalamus co-migrated exactly with synthetic α-MSH, whereas the arcuate nucleus contained 5 peptides cross-reacting with α-MSH antibodies, 4 of them being different from standard α-MSH. Significant amounts of biologically active melanotropin, which migrated on Sephadex G-25 columns like synthetic α-MSH, were also detected in both the arcuate nucleus and dorsolateral hypothalamus. Taken together, these data demonstrate that the α-MSH cell bodies located in the dorsolateral hypothalamus specifically produce authentic α-MSH, whereas the α-MSH cell bodies in the arcuate nucleus also contain ACTH, β-endorphin and several peptides immunologically related but not identical to α-MSH.

Amniotic fluid β-endorphin and α-melanocyte-stimulating hormone immunoreactivity in normal and complicated pregnancies

β-Endorphin (β-EP) and α-melanocyte-stimulating hormone (α-MSH) are members of a family of peptides which are found in the intermediate lobe of the fetal pituitary gland and placenta. In the present study, concentrations of immunoreactive β-EP (iβ-EP) and α-MSH were determined by radioimmunoassay in the amniotic fluid compartment of both normal (n = 72) and complicated (n = 44) pregnancies. In normal pregnancies, there was a significant (p < 0.05) fall in the man amniotic fluid iβ-EP from the second to third trimester (from 175 to 132 pg/ml). A similar, but not statistically significant, decline in the mean amniotic fluid concentrations of α-MSH (from 46 to 34 pg/ml) was also noted. Concentrations of amniotic fluid iβ-EP showed a significant correlation with α-MSH (r = 0.61, p < 0.001). Significant elevations of mean amniotic fluid iβ-EP but not α-MSH were found in pregnancies complicated by intrauterine growth retardation (215 pg/ml, p < 0.0001) and premature labor (225 pg/ml, p < 0.0001). Our findings suggest that levels of iβ-EP and α-MSH in amniotic fluid decrease from the second to the third trimester of pregnancy, and that elevation in iβ-EP concentration in amniotic fluid, in all probability, reflects the secretion of β-EP by the fetus in response to fetal distress.

Effects of chronic lithium, clorgyline, imipramine, fluphenazine and constant darkness on the α-melanotropin content and circadian rhythm in rat brain

The effects of constant darkness, chronic lithium, clorgyline, imipramine and fluphenazine treatment on the content and diurnal rhythm of α-MSH in rat forebrain were investigated. The persistence of the α-MSH rhythm in constant darkness demonstrated that the rhythm was circadian in nature. Constant darkness increased the 24 h mean α-MSH concentration in brain while lithium, fluphenazine and imipramine decreased it. In addition, imiparamine and clorgyline delayed the phase of the α-MSH circadian rhythm while lithium advanced it.

Changes in α-melanotropin in discrete brain areas of isolated aggressive mice

Adult male Swiss-Webster (NIH) mice were isolated for 6 weeks. Aggressive behavior was tested on 2 occasions, 24 hours apart. Immediately following the 2nd test period, aggressors and isolated non-aggressors (controls) were decapitated and α-MSH concentration was measured in discrete areas of the brain. Only the nucleus accumbens and preoptic lateralis of the aggressors, showed a higher level of α-MSH when compared to the controls. The significance of these changes is discussed.

Dopaminergic control of aldosterone: Modulation of the response of rat adrenal zona glomerulosa cells to α-MSH by pretreatment with bromocriptine or metoclopramide

Bromocriptine treatment in rats (3 mg/kg per day, 7 days) significantly reduced α-MSH and aldosterone plasma levels 2 hrs after the final treatment in animals on low, normal and high sodium diets. α-MSH dose response curves for corticosterone and 18-hydroxydeoxycorticosterone (18-OH-DOC) in subsequently incubated glomerulosa cells gave stimulation at lower concentrations of α-MSH (10 −10 moles per litre) than in cells from untreated animals (10 −9 moles per 1). Curves for aldosterone (ald) and 18-hydroxycorticosterone (18-OH-B) were also affected in cells from animals on a low sodium diet. Fasciculata-reticularis cell responses to ACTH were unaffected. Metoclopramide (4 mg/kg per day, 7 days) elevated plasma α-MSH, although ald was unaffected, but inhibited the glomerulosa cell response to α-MSH in vitro . Acute dopaminergic responses in plasma ald may be mediated through α-MSH in rats, but chronically α-MSH may down-regulate glomerulosa cell α-MSH receptors. It is unlikely that α-MSH mediates the adrenocortical response to sodium depletion.

Distribution of an enzyme which acetylates α-melanocyte stimulating hormone in rat brain and pituitary gland and effects of arcuate nucleus lesions

This study describes the distribution of an α-melanocyte stimulating hormone (α-MSH) acetyltransferase (MAT) in rat brain and pituitary gland. Highest activities of MAT were found in the neurointermediate lobe of the pituitary gland with the anterior lobe containing slightly less. Within the brain, lowest MAT activities were measured in the hypothalamus, the region which contained the highest concentrations of α-MSH. Relatively high enzyme activities of MAT were measured in the hippocampus, cortex and cerebellum—regions with very low α-MSH concentrations. The fact that MAT activity levels did not parallel α-MSH concentrations indicates that MAT was not solely localized to α-MSH synthesizing neurons or endocrine cells. Furthermore, arcuate nucleus lesions which depleted brain α-MSH failed to deplete MAT activity. Although MAT was not solely localized to α-MSH synthesizing cells, it may have functional significance for α-MSH acetylation due to compartmentalization with α-MSH in α-MSH synthesizing endocrine cells and neurons. Alternatively a second regionally specific MAT may exist.

Responses of rat fetal adrenals to synthetic adrenocorticotrophic hormone and alpha-melanocyte-stimulating hormone in-vivo and in-vitro studies.

The response of the adrenals from rat fetuses at 16, 18 and 20 days of gestation to 1-24 ACTH and alpha-melanocyte-stimulating hormone (alpha-MSH) was studied in vitro. The response to 1-24 ACTH increased as gestation progressed. By the end of fetal life, corticosterone release induced by ACTH from whole adrenals was greater than that observed with adrenal tissue from non-pregnant adult female rats. High doses of alpha-MSH also stimulated adrenal activity but the response to ACTH was always higher than that to alpha-MSH. The effect of 1-24 ACTH and alpha-MSH on fetal adrenal growth was also compared in vivo. The adrenal atrophy induced by fetal hypophysectomy on day 17 of gestation could be prevented by i.m. administration of 10 microgram 1-24 ACTH or alpha-MSH. However, the adrenal growth was greater in ACTH-treated fetuses than in alpha-MSH treated ones. Later in gestation, between days 19 and 20, 1-24 ACTH but not alpha-MSH was able to prevent atrophy induced by fetal hypophysectomy. These findings are discussed in relation to the literature on levels of ACTH and alpha-MSH in the plasma and pituitary glands of the rat throughout the last third of gestation. High levels of ACTH in the fetal circulation contrast sharply with very weak or undetectable concentrations of alpha-MSH. Since the present data suggest that both trophic and steroidogenic activities of ACTH were greater than those of alpha-MSH, it may be concluded that ACTH but not alpha-MSH plays a major physiological role during gestation in the regulation of both fetal adrenal growth and function in the rat.

The effect of fever on central α-MSH concentrations in the rabbit

The recent demonstration of the ability of centrally administered alpha-MSH to reduce pyrogen-induced fever in rabbits suggested a role for the peptide in central temperature control. Using a sensitive radioimmunoassay we have established the presence of immunoreactive alpha-MSH (ir alpha-MSH) in discrete CNS sites of the rabbit and have demonstrated a distribution of this activity similar to that seen in other species. Pyrogen-induced fever did not alter concentrations of ir alpha-MSH in tissue extracted from the paraventricular region of the hypothalamus, median eminence, midbrain central grey, preoptic/anterior hypothalamic region, pineal or pituitary. A decrease in arcuate nucleus ir alpha-MSH content was seen in febrile animals, although this change was not statistically significant. Significantly higher levels of ir alpha-MSH were detected in septal extracts of febrile rabbits (0.81/+-0.10 pg/micrograms protein, n=9) than in extracts from afebrile controls (0.54/+-0.04, n=17). This finding suggests the septum to be the central site of action of alpha-MSH to inhibit fever and further implies a modulatory role of the peptide in the central control of thermoregulation.

Changes in plasma, pituitary and brain α-MSH content in rats from birth to sexual maturity

Immunoreactive α-MSH was measured in plasma, pituitary and brain of male and female rats on the day of birth and at intervals afterwards up to 55–70 days of age. Plasma α-MSH concentrations on the day of birth were 528 ± 111 pg/ml and 406 ± 137 pg/ml in famel and male rats, respectively. Plasma α-MSH concentrations then fell and remained low until the onset of sexual maturity when they again rose reaching 406 ± 38 pg/ml in 70 day old females and 312 ± 46 pg/ml in 55 day old males. Pituitary α-MSH concentrations also changed with age and in male rats generally reflected the changes in plasma α-MSH concentrations. In females, on the other hand, pituitary α-MSH concentrations showed a gradual increase with age. Concentrations of α-MSH in the hypothalamus and brain increased with age and as with plasma and pituitary reached peak values in sexually mature animals. These findings are consistent with the idea that both piutiary and brain α-MSH have a role in sexually mature rats. Although α-MSH may have a role in sexual behaviour there was no evidence of any change in brain α-MSH throughout the estrous cycle.

Demonstration of an endogenous circadian rhythm of α-melanocyte stimulating hormone in the rat pineal gland

α-Melanocyte stimulating hormone (α-MSH) has been identified and characterized in the rat pineal gland by a combination of immunochemical and high pressure liquid chromatographic techniques. The immunoreactivity in pineal extracts was separated into two chromatographic components. The major component had a retention time identical to that of α-MSH while the minor component eluted just slightly before standard α-MSH. Male rats maintained in a 12 h photoperiod demonstrated a marked circadian rhythm in pineal α-MSH concentration. Concentrations that peaked at 07.00 h, 1 h after the lights were turned on, were greater than 5 times the nadir which occurred at 01.00 h. Animals housed in chronic dark for 7 days maintained the diurnal variation of α-MSH concentrations. However, in chronically dark housed rats, the peak shifted to 05.00 h and was greater than 10 times the nadir of this rhythm and approximately 4 times the peak at 07.00 in alternating light/dark conditions. Rats exposed to chronic light for 7 days maintained a pineal α-MSH rhythm although the amplitude of the peak was significantly decreased compared to the rhythm in animals housed in alternating light/dark conditions. Neither hypophysectomy nor superior cervical ganglionectomy had any effect on the α-MSH rhythm. Lesion of the arcuate nucleus, the major source of α-MSH-containing nerves in the brain, did not significantly affect pineal α-MSH concentrations. These data demonstrate a circadian α-MSH rhythm in the rat pineal and suggest an α-MSH involvement in the rhythmic processes of the pineal gland.

Identification and distribution of α-melanotropin in discrete regions of the cat brain

The distribution of α-melantropin-like material (α-MSH) was determined in rat brain using a microdissection technique combined with radioimmunoassay. Highest concentrations of α-MSH were noted in the hypothalamus and preoptic area. Particularly high concentrations were observed in the arcuate nucleus, median eminence, suprachiasmatic nucleus, periventricular nucleus and the medial preoptic nucleus. The thalamic paraventricular nucleus also contained high α-MSH concentrations. Moderate to low concentrations were noted in other thalamic, septal, amygdaloid and midbrain nuclei. Low concentrations were observed in cortical and striatal regions.

Identification, characterization and stereotaxic mapping of intraneuronal α-melanocyte stimulating hormone-like immunoreactive peptides in discrete regions of the rat brain

A highly specific antibody to α-melanocyte stimulating hormone (α-MSH) was used to histochemically localize and biochemically identify and quantitate α-MSH immunoreactivity in nerve fibers and cell bodies of the rat brain. α-MSH-like immunoreactivity was contained in fibers throughout the brain. The distribution of α-MSH was determined by immunocytochemistry as well as by radioimmunoassay combined with microdissection techniques. High concentrations of α-MSH were contained in the nucleus interstitialis stria terminalis, the median eminence and the medial preoptic, anterior hypothalamic, periventricular, paraventricular, arcuate, dorsomedial, and posterior hypothalamic nuclei. Moderate α-MSH concentrations were noted in the amygdala, septum, central gray, dorsal raphe, and the nucleus tractus solitarius. Cell bodies containing α-MSH were observed only in the arcuate nucleus. The α-MSH-like compound in brain had similar immunochemical and electrophoretic properties of standard α-MSH but high pressure liquid chromatographic analysis demonstrated that the α-MSH-like immunoreactivity was comprised of one major and two minor components. The major immunoreactive peak had an identical retention time as α-MSH and therefore may be chemically identical to α-MSH. The similar retention times and immunoreactivity of the other two compounds suggest a similarity in size and structure to α-MSH. These observations demonstrate that fibers containing α-MSH emanate from the arcuate nucleus to innervate many other regions of the rat brain.

Subcellular localization of immunoreactive α-melanocyte stimulating hormone in human brain

The regional and subcellular distribution of immunoreactive α-melanocyte stimulating hormone (α-MSH 1) in the post mortem adult human brain was investigated. α-MSH i was highly concentrated in medial basal hypothalamic tissue (1.02 ng/mg protein). Lower levels of α-MSH i were present in the optic chiasm and mammillary bodies, 0.08 and 0.11 ng/mg protein, respectively. The concentrations of α-MSH i in cerebellum and frontal cerebral cortex were 1/1,000th that of the medial basal hypothalamus. When medial basal hypothalamic homogenates were subjected to discontinuous or continuous sucrose density gradients, α-MSH i was found to be associated primarily with subcellular particles which resembled isolated nerve terminals, i.e., synaptosomes. Low to undetectable amounts of α-MSH i were found in the cytosol or the myelin/microsome fraction of the gradients. The results of these studies are consistent with the view that α-MSH is a neuronal peptide in the human brain.

Hypothalamic-Pituitary Vasculature: Evidence for Retrograde Blood Flow in the Pituitary Stalk1

Retrograde transport of pituitary hormones in the pituitary stalk vasculature was investigated in anesthetized male rats in which the pituitary gland was intact and in animals in which the anterior pituitary, posterior pituitary, or entire pituitary gland had been removed 30 to 60 min before use. Blood was collected for 1.5 to 2 h by free flow from a single long portal vessel through a microcannula, the tip of which was pointed toward the hypothalamus. An arterial blood sample was obtained at the end of each collection of portal blood. The concentrations (ng\ml) of LH, TSH, prolactin, ACTH, α-MSH, and vasopressin, determined by radioimmunoassay, in portal plasma from rats with intact pituitary glands were as follows: LH, 2,320 ± 874 (mean and SE); TSH, 10,180 ± 1,471; prolactin, 4,858 ± 884; ACTH, 82 ± 17.0; a-MSH, 103 ± 17.8; vasopressin, 2.4 ± 1.0. The concentrations of these hormones in arterial plasma of these rats were as follows: LH, <20; TSH, 149 ± 22; prolactin, 25 ± 5.0; ACTH, 0.36 ± 0.05; α-MSH,...