Role of alpha-MSH and related peptides in the central nervous system

Alpha-melanocyte stimulating hormone (alpha-MSH) is a tridecapeptide secreted by intermediate lobe cells and synthesized in the brain as well. As a hormonal peptide, the physiological function of alpha-MSH consists mainly in the control of pigment movements within dermal melanophores. At the pituitary level, alpha-MSH secretion is under multifactorial control: it is inhibited by dopamine and GABA and stimulated by corticoliberin (CRF), thyroliberin (TRH), beta-adrenergic agonists and (or) serotonin. Identification of alpha-MSH containing neurons in the hypothalamus and other brain regions (septum, thalamus, mid-brain, striatum, hippocampus, cerebral cortex and spinal cord) has been carried out by means of immunological and biochemical techniques combined with bioassays. In the central nervous system (CNS) as in the hypophysis, alpha-MSH is synthesized from a high molecular weight precursor, pro-opiomelanocortin (POMC). Maturation of this protein yield similar end products in the hypothalamus and the intermediate lobe. Several peptides chemically related to alpha-MSH are generated including the desacetyl and monoacetyl (authentic alpha-MSH) forms; the latter has the greatest behavioral activity. The demonstration that alpha-MSH has numerous central nervous system effects has raised the possibility that this neuropeptide acts as a neuromodulator or a neurotransmitter. In the rat, intra-cerebroventricular administration of ACTH/MSH peptides induces the stretching-yawning syndrome (SYS) which is frequently preceded by excessive grooming. This excessive grooming is blocked by neuroleptics indicating that the central dopaminergic neurons are implicated in this behavioral effect of the peptide. alpha-MSH is involved in memory, arousal and attention; in hypophysectomized animals, the learning ability is restored after administration of MSH or related peptides. Injection of alpha-MSH delays also extinction of passive avoidance behavior and affects performances motivated by hunger as well as aggressive behavior. Recent studies concerning the role of alpha-MSH have been undertaken in human beings. The effects of MSH-related peptides favour a role of these peptides in arousal: they maintain a high level of vigilance and improve visual discrimination. These behavioral changes were accompanied by marked changes in CNS electrophysiology. Current studies, which aim at establishing a neurotransmitter function for alpha-MSH, concern the distribution and characterization of alpha-MSH receptors in the central nervous system and the mechanism controlling the release of neuronal alpha-MSH.     

Behavioral influences of the neuropeptides ACTH and MSH: A methodological review

During the past 20 years, there has been repidly accumulating evidence that polypeptide hormones of pituitary origin are important mediators of behavior. The research has been focused primarily upon the actions of adrenocorticotropic hormone (ACTH) and melanocyte-stimulating hormone (MSH). A large number of methodological difficulties have developed and are in need of careful review. Important pharmacological and behavioral variables are reviewed both in general and as they relate to current research on the actions of ACTH and MSH. In addition, suggestions are given for control and/or study of these variables. Finally, the main integrating hypotheses, fear, memory and attention, are reviewed and evaluated.     

Melanocortins stimulate proliferation and induce morphological changes in cultured rat astrocytes by distinct transducing mechanisms.

Melanocyte stimulating hormone (MSH), adrenocorticotropic hormone (ACTH), and several peptides derived from pro-opiomelanocortin, are present in the dorsolateral hypothalamus and arcuate nucleus of several vertebrate species. These peptides affect central nervous system (CNS) functions including behavior, memory, and foetal brain development. In this study we investigated the effects of ACTH1-24, ACTH1-17, ACTH4-10, alpha-MSH, beta-MSH, and a potent analog (Nle4,D-Phe7)-alpha-MSH (melanocortins) on immunocytochemically defined astroglial cells prepared from primary cultures of 1-2-day-old rat brains. A cyclic adenosine 3',5'-monophosphate (cAMP) response to the melanocortins was only detected in astrocytes and not in other cell types in the culture. The extent of the cAMP response was greatest on day 21, the latest time tested. On the other hand, (methyl3H)-thymidine incorporation in astrocytes was significantly stimulated (1.5-2-fold) by melanocortins only in 7 and not in 14 and 21 day cultures. This mitogenic activity of melanocortins was not mimicked by other agents such as forskolin or isoproterenol which efficiently stimulate cAMP production in astrocytes. ACTH1-17 as a melanocortin representative induced significant morphological changes in 7 and 14 day cultures which included rounding of the cell body and process extension. This response, however, resembled that induced by forskolin and hence appears to be cAMP mediated. These findings suggest that astrocytes in the CNS may serve as a target for melanocortins. These peptides appear to affect differentiation and proliferation of these cells during certain developmental periods. While the morphological effects of melanocortins seem to be cAMP mediated, induction of proliferation of the astrocytes by melanocortins appears to involve an alternative signal transduction pathway.     

Melanotropin receptors in the brain are differentially distributed and recognize both corticotropin and alpha-melanocyte stimulating hormone

Melanotropinergic neurons in the brain may mediate the known modulatory effects of alpha-melanocyte stimulating hormone (alpha-MSH) and adrenocorticotropic hormone (ACTH) on brain functions including thermoregulation, pituitary regulation, learning and behavior. To determine the distribution of brain melanotropin receptors, we used biologically active [125I]Nle4, D-Phe7-alpha-MSH ([125I]NDP-MSH) for in situ binding and autoradiography in frozen rat brain sections. Specific (alpha-MSH-inhibitable) [125I]NDP-MSH binding sites were distributed in a region-specific pattern, and were present in numerous structures within the septal area, hypothalamus, thalamus, epithalamus, olfactostriatal complex, and midbrain. Each brain structure studied showed a characteristic, reproducible distribution and relative intensity of binding. Receptor peptide selectivity was assessed by comparing the dose-response relationships for inhibition of binding by alpha-MSH, NDP-MSH and ACTH. In all brain structures studied, the 3 peptides gave comparable maximal inhibition of tracer binding, indicating that all detectable binding sites recognized all 3 melanotropins. The respective relative potencies were: NDP-MSH (EC50 = 1.7 +/- 0.6 nM) greater than alpha-MSH (EC50 = 46.9 +/- 11.7 nM) = ACTH. These results provide a preliminary neuroanatomic map of potential target sites for melanotropin actions, and indicate that these sites are capable of recognizing multiple products of the intrinsic melanotropinergic system of the brain.     

Ingested (oral) alpha-MSH inhibits acute EAE

Ingested type I IFN and SIRS peptide administered orally inhibit EAE. We examined whether another immunoactive protein, tridecapeptide alpha-MSH, would have similar anti-inflammatory effects in EAE after oral administration. B6 mice were immunized with MOG peptide 35-55 and gavaged with 0.1 ml of control saline or alpha-MSH peptide starting on day -7 preceding active immunization, and continuing through day +14 post-immunization. Alpha-MSH peptide delayed disease onset and decreased inflammatory foci. CNS lymphocytes showed decreases in Th1-like encephalitogenic cytokines IL-2 and IL12p70 in the alpha-MSH fed group compared to the mock fed group. For Th2-like counter-regulatory cytokines, there were increases in peripheral SDF-1 levels comparing alpha-MSH fed vs mock fed groups. There were decreases of chemokines MIP-1-alpha and MIP-1-gamma in the CNS comparing alpha-MSH fed mice vs mock fed mice. Ingested (orally administered) alpha-MSH peptide can reduce clinical disease and inhibit CNS inflammation by decreasing migration of antigen driven CNS Th1 cells into the target organ.     

Dexamethasone facilitates release of the neuropeptide alpha-MSH.

alpha-Melanocyte stimulating hormone (alpha-MSH) has important host defense properties, in part similar to those of corticosteroids. Previous research suggests that secretion of alpha-MSH and of ACTH are controlled separately. The relationship between release of alpha-MSH and the activity of the hypothalamic-pituitary-adrenal axis in the rabbit was examined by monitoring changes in circulating alpha-MSH, ACTH, and corticosterone in response to endotoxin and corticotropin-releasing hormone (CRH), both with and without dexamethasone pretreatment. Endotoxin (1 microgram/kg IV) did not cause alpha-MSH release, but it did increase plasma concentrations of ACTH and corticosterone. Similarly, CRH (1 and 10 micrograms/kg IV) did not affect plasma alpha-MSH, whereas it stimulated ACTH and corticosterone release. Dexamethasone pretreatment abolished the responses of ACTH and corticosterone to either stimulus and did not modify circulating alpha-MSH after CRH. In contrast, dexamethasone pretreatment did result in a significant increase in plasma alpha-MSH after a dose of endotoxin that was ineffective alone. These data indicate that corticosteroids can facilitate the release of alpha-MSH, a powerful anti-inflammatory hormone. Since corticosteroids are released with certain challenges, this facilitatory activity may be important to the host response.     

Isolation and structural characterization of peptides related to alpha- and gamma-melanocyte-stimulating hormone (MSH) from the frog brain.

Peptides that are derived from the processing of proopiomelanocortin were isolated in pure form from the brain of the frog Rana ridibunda. The primary structure of the most abundant of those peptides was established as: Ser-Tyr-Ser-Met-Glu-His-Phe-Arg-Trp-Gly-Lys-Pro-Val. This amino acid sequence is identical to that of mammalian and frog pituitary alpha-melanocyte-stimulating hormone (MSH) and the peptide co-eluted with synthetic desacetyl alpha-MSH, indicating that it is COOH-terminally alpha-amidated. A second component, which exhibited a shorter retention time, co-eluted with the glycine-extended form of desacetyl alpha-MSH [ACTH(1-14)]. The primary structure of the third peptide isolated in pure form from the brain extract was established as: Lys-Tyr-Val-Met-Ser-His-Phe-Arg-Trp-Asn-Lys-Phe-NH2. This sequence corresponds to Lys-gamma 1-MSH as predicted from the nucleotide sequence of frog proopiomelanocortin. The presence of substantial amounts of desacetyl alpha-MSH and Lys-gamma 1-MSH in the frog brain supports the concept that, in amphibia, melanotropins may act as neurotransmitters and/or neuromodulators as well as hormonal peptides.     

Central actions of CRF on thermogenesis are mediated by pro-opiomelanocortin products

Corticotrophin-releasing factor (CRF) causes central activation of thermogenesis. The aim of this study was to investigate whether this action is mediated by ACTH or other peptides derived from the ACTH precursor pro-opiomelanocortin (POMC) within the CNS. Central (intracerebroventricular) injection of rat CRF caused dose-dependent increases in resting oxygen consumption (VO2) in conscious rats (maximal 26 +/- 5% at 2 nmol CRF). These responses were significantly attenuated by pretreatment (i.c.v.) with either a monoclonal antibody raised to gamma 1MSH or with naloxone which antagonises beta-endorphin (beta-EP) actions. The increases were not affected by pretreatment with monoclonal antibodies to ACTH or the N-terminal of POMC. Central injections of gamma 1-melanocyte-stimulating hormone (MSH) or beta-EP caused dose-dependent increases in VO2 (maximal at 0.5-1.5 pmol) and these were markedly inhibited by pretreatment with the anti-gamma 1-MSH antibody or naloxone respectively. Injection of ACTH or alpha MSH did not significantly affect VO2 at doses up to 2 nmol. These data indicate that the central actions of CRF on thermogenesis may be mediated, at least in part, by release of gamma MSH and/or beta-EP.     

Effects of ACTH4−10 on vestibular compensation

ACTH4-10, a fragment of the adrenocorticotropic hormone (ACTH) molecule, has marked effects on the compensation process following unilateral labyrinthectomy. In Rana temporaria ACTH4-10-treatment (5-250 micrograms/kg) influences both the acquisition and the maintenance of the compensated state. The compensation process is slowed down by hypophysectomy but can then be restored by the administration of ACTH4-10. It is concluded that ACTH-like neuropeptides might physiologically be involved in the plastic processes underlying functional recovery from CNS lesions.     

Efficacy of ACTH 4–9 analog, methylphenidate, and placebo on attention deficit disorder with hyperkinesis

1. The present study compared the efficacy of ACTH 4–9 analog, methylphenidate, and placebo on attention deficit disorder with 30 (HK) children with attention deficit disorder and psychometric measurements.

2. HK children were randomly placed into three groups, according to body weight, and treated with placebo, methylphenidate, and adrenocorticotropic hormone fractions (ACTH 4–9 analog).

3. The results revealed that HK children on methylphenidate manifested a significantly greater vasomotor reactivity, behavioral improvement, and learning receptivity than HK children taking ACTH 4–9 analog and/or placebo.

4. Future research implications with ACTH 4–9 and HK children are discussed.     

Characterization of immunoreactive alpha-melanocyte stimulating hormone (alpha-MSHi) in human brain tissue

Adult human hypothalamic tissue was analyzed for the presence of products generated by post-cleavage processing of ACTH 1-13. The displacement curve generated by immunoreactive alpha-melanocyte stimulating hormone (alpha-MSHi) in extracts (5 M acetic acid or acidified ethanol) of adult human hypothalamic tissue was parallel to the alpha-MSH radioimmunoassay standard curve, and alpha-MSHi eluted as a single peak on Sephadex G-10, G-25, and G-50 columns in a manner identical to that of synthetic alpha-MSH. The alpha-MSHi was clearly distinguishable in its mobility on Sephadex G-50 columns from such peptides as beta-MSH, luteinizing hormone releasing hormone, and ACTH. After high performance liquid chromatography of extracts of hypothalamic tissue from young as well as aged men and women, we found that the major peak of alpha-MSHi (75-95% of total alpha-MSHi) coeluted with desacetyl alpha-MSH (ACTH 1-13 amide) rather than alpha-MSH. We suggest that desacetyl alpha-MSH, rather than alpha-MSH, is the predominant alpha-MSHi in adult human brain tissue.     

Effect of stress-related hormones on short term memory

Adrenocorticotropic hormone (ACTH)1-24, ACTH4-10, corticosterone (CS) or arginine vasopressin (AVP) was administered subcutaneously to one day-old chicks immediately after learning a single trial passive avoidance task. Chicks were pretreated with 2 mM KC1 or 4 mM monosodium glutamate 5 min before learning. With KC1 or monosodium glutamate alone, no evidence of memory was observed on retention tests carried out as early as 5 min and as late as 24 h postlearning. However, the addition of ACTH1-24, ACTH4-10 or AVP to KC1-pretreated animals yielded normal retention levels up till 10 min, 10 min and 20 min after learning, respectively. Similar results were obtained with ACTH1-24 and AVP given to glutamate-pretreated birds. CS had no effect on KC1- or glutamate-induced amnesia. The calcium channel blocker, lanthanum chloride, also inhibited the formation of short-term memory, with amnesia still present as late as 24 h following learning. ACTH1-24, but not CS or AVP, yielded normal retention levels until 10 min postlearning in the presence of lanthanum chloride. Thus ACTH1-24 and AVP can overcome KC1 or glutamate inhibition of STM formation but will not prevent subsequent amnesia. The mechanisms underlying this action of ACTH1-24 and AVP are different. The possibility that the effect of ACTH1-24 is related to the role of calcium in STM formation is explored.     

Adrenocorticotrophic hormone and melanocyte stimulating hormone do not affect performance in hippocampus-lesioned or control pigeons

Pigeons, given bilateral lesions of the hippocampus, as well as unoperated control subjects, were trained on an operant task in which they had to peck a left hand key if both members of a successively presented stimulus pair were the same, and a right hand key if they were different. During extensive training, the 'same' pairs (green-green or red-red) were presented at twice the frequency of the 'different' pairs (green-red or red-green), thus biasing responding to the left hand key. The effects of intramuscular (i.m.) injections (0, 10 and 100 micrograms/kg) of an active fragment of adrenocorticotrophic hormone (ACTH4-10) and alpha-melanocyte stimulating hormone (MSH) were then studied. On any given test (peptide or saline) day, all stimulus pairs were presented with equal frequency. Although the control birds' performance was consistently better than the lesioned birds, neither ACTH4-10 nor MSH affected performance under any conditions. It is concluded that peripheral injection of these peptides has little, if any, effect on cognitive behaviour in birds.     

Endogenous alpha-MSH modulates the hypothalamic-pituitary-adrenal response to the cytokine interleukin-1beta

We and others have previously shown that exogenous alpha-MSH antagonizes the stimulatory effects of the cytokine interleukin (IL)-1 on the hypothalamic-pituitary-adrenal (HPA) axis. It is currently unknown, however, if endogenous alpha-MSH plays a physiological role in regulating the HPA response to IL-1. We have therefore examined the HPA response to IL-1beta in rats pretreated with an affinity purified alpha-MSH antiserum (AS) infused intracerebroventricularly to neutralize endogenous alpha-MSH within the brain. alpha-MSH AS or a similarly purified fraction of normal rabbit serum (NRS) was injected intracerebroventricularly at 16 h and at 1 h prior to the i.c.v. injection of IL-1beta (2 ng or 20 ng) and blood samples were collected through an indwelling atrial catheter. After 2 ng IL-1beta, the adrenocorticotropic hormone (ACTH) response was significantly greater in the alpha-MSH AS treated rats (n = 7) compared to the NRS treated rats (n = 7) (P <0.01); the mean ACTH level rose to a peak of 594+208 pg/ml in the alpha-MSH AS treated rats vs 274+/-122 pg/ml in the NRS treated rats. The area under the ACTH response curve in the alpha-MSH AS treated animals was 181% of that in the NRS treated animals (P<0.05). A significant effect of alpha-MSH AS on the corticosterone response to i.c.v. IL-1beta was also noted during the first 3 h of the study (P<0.05). The mean area under the corticosterone response curve for the first 3 h in the alpha-MSH AS treated animals was 144% of that in the NRS treated animals (P <0.05). After 20 ng IL-1beta, the ACTH response over time was again significantly greater in the alpha-MSH AS treated rats (n=8) compared to the NRS treated rats (n=9) (P<0.02); the mean ACTH level rose to a peak of 673+/-190 pg/ml after alpha-MSH AS vs 490+/-115 pg/ml after NRS. Corticosterone levels rose to a peak of 42+/-3.9 microg/dl in the alpha-MSH AS treated rats vs 37+/-4.6 microg/dl in the NRS treated rats; this difference was not significant. We conclude that the IL-1beta induced stimulation of ACTH is significantly enhanced by antagonizing the activity of alpha-MSH. These results support a physiological role for endogenous alpha-MSH in limiting the HPA response to this inflammatory cytokine.     

Beneficial effect of chronic treatment with Org 2766 and alpha-MSH on impaired reversal learning of rats with bilateral lesions of the parafascicular area

The effects of chronic treatment with the ACTH-(4-9) analogue Org 2766, alpha-MSH, and gamma 2-MSH were studied on T-maze reversal learning and on behavior assessed on the basis of open-field and other gross behavioral activities, grasping responses, inspection of various reflexes and electrical footshock sensitivity of rats with parafascicular lesions or sham-lesions. Repeated administration of Org 2766 and alpha-MSH to parafascicular area-lesioned rats resulted in functional recovery of impaired T-maze reversal learning. The structurally related neuropeptide gamma 2-MSH was without any effect. The alpha-MSH effect did not depend on time after lesioning as treatments during the first or second post-operative week were equally effective. Chronic peptide treatments did not change disturbed motor functions of parafascicular-lesioned rats, as measured by open-field activity, other gross behavioral activities and grasping responses. Since acute peptide treatments did not affect the impaired reversal learning performance of lesioned rats, the beneficial effect of Org 2766 and alpha-MSH could not be explained as a short-term effect on attention and motivation. It was more likely to be an accelerated recovery of cognitive function as a result of long-term neurotropic influences.     

Acetylation alters the feeding response to MSH and beta-endorphin

The effects on food intake of the N-acetylation of MSH and beta-endorphin have been examined following their injection into the third ventricle. Desacetyl-MSH and alpha-MSH were injected into fasted rats, and beta-endorphin and N-acetyl-beta-endorphin into fed rats. Desacetyl-MSH had no effect on food intake following ICV injection into food-deprived rats at any dose between 100 and 2500 pmoles. Alpha-MSH, the N-acetylated form of MSH, on the other hand, showed a highly significant inhibition of food intake in food-deprived rats with doses of 100 and 250 pmoles but no effect with the higher doses. With beta-endorphin, there was a dose-related biphasic effect. One hour after injection of beta-endorphin (2500 pmole) food intake was inhibited whereas the lowest dose, 100 pmole, significantly stimulated it. By 3 hours, the 2 lowest doses of beta-endorphin both significantly stimulated food intake, but the highest dose remained inhibitory. By 6 hours all doses of beta-endorphin stimulated food intake compared to the vehicle-treated animals. In contrast, N-acetylation of beta-endorphin eliminated all effects on food intake following injection into the third ventricle. These data suggest that N-acetylation of products formed by the processing of POMC can markedly alter their biological properties.     

Plasma levels of alpha-melanotropin and ACTH-like immunoreactivities do not vary by season or skin type in women from southern and central Europe

Melanotropins (MSH) are involved in tanning by stimulating melanocytes via the activation of the melanocortin-1 receptor to melanin production. Its main site of production is the pituitary gland, but alpha-MSH and related ACTH peptides are produced at other sites, including the skin. It has been hypothesized that systemic levels of alpha-MSH are controlled by a varying UV radiation (UVR) exposure. A seasonal rhythm of plasma levels has been proposed by some authors. We investigated healthy females in southern Spain and central Austria in summer and winter. The alpha-MSH and ACTH-like immunoreactivity plasma levels did not present marked differences between the groups of Malaga and Linz, dark and light skin and between seasons. An association of alpha-MSH to ACTH or cortisol levels could not be observed. Individual values of alpha-MSH were shown to be relatively constant at both times of measurement (p<0.05 by rank correlation) indicating an independent personal disposition for individual systemic alpha-MSH immunoreactivity levels. Our data do not support the concept of a marked involvement of melanotropins of pituitary origin in tanning, and no seasonal rhythm was observed.     

Effects of preoptic microinjections of alpha-MSH on fever and normal temperature control in rabbits

alpha-MSH within the septal region of the brain has been implicated in fever control; this peptide and ACTH (1-24), which contains the alpha-MSH amino acid sequence, reduce fever when given intracerebroventricularly (ICV) or peripherally. These peptides also cause hypothermia when given in doses larger than those required to reduce fever. Both peptides occur naturally within the preoptic PO region of the brain, the CNS locus of primary temperature control. alpha-MSH (350 ng) injected bilaterally into the PO region via chronic cannulas reduced fever caused in six rabbits by IV injection of IL-1 (interleukin 1, endogenous or leukocyte pyrogen) but had no effect in afebrile animals. A larger dose (1.5 micrograms) not only reduced fever but caused hypothermia in 12 rabbits. In separate experiments PO injections of ACTH (1-24) (1 microgram) reduced normal temperature. In the same six rabbits alpha-MSH (1 microgram) caused slightly smaller hypothermia. alpha-MSH (1.5 micrograms) also had no effect in 8 afebrile rabbits when injected into the septum. The primary conclusion is that alpha-MSH receptors within the PO region can contribute to both the antipyretic and hypothermic actions that are observed after ICV and peripheral administration of the peptide.     

ACTH- and alpha-MSH-induced grooming, stretching, yawning and penile erection in male rats: site of action in the brain and role of melanocortin receptors

The effect of adrenocorticotropin (ACTH)(1-24) and alpha-melanocyte stimulating hormone (alpha-MSH) on grooming, stretching, yawning and penile erection was studied after injection into different brain areas. Both peptides induce the above responses when injected into the hypothalamic periventricular region of the third ventricle. This region includes the paraventricular nucleus, the dorsomedial nucleus, the ventromedial nucleus and the anterior hypothalamic area. The minimal effective dose of both peptides was 0.5 microg and the maximal effect was seen with 2 microg, the highest dose tested. Irrespective of the injection site, grooming started 5-7 min after injection of either peptide, while stretching, yawning and penile erection started only after 15-35 min and lasted for 90-120 min. In contrast both peptides were ineffective when injected into the preoptic area, the caudate nucleus or the CA1 field of the hippocampus. Grooming, stretching and yawning, but not penile erection, were prevented by cyclic[AcCys(11), D-Nal(14), Cys(18), AspNH(2)(22)]-beta-MSH (11-22) (HS014), a selective melanocortin 4 receptor antagonist, injected into the same periventricular area 10 min before of ACTH(1-24) or alpha-MSH. The results show that ACTH(1-24) and alpha-MSH act in the hypothalamic periventricular region to induce the above responses and that grooming, stretching and yawning, but not penile erection, are mediated by melanocortin 4 receptors.