Search

Melanocortins are proopiomelanocortin-derived peptides that include adrenocorticotropic hormone [ACTH (1-39)], alpha-melanocyte-stimulating hormone [alpha-MSH (1-13)], and related amino acid sequences. Melanocortin peptides have potent antiinflammatory/anticytokine activity. Because cytokines such as interleukin 1 (IL-1) and tumor necrosis factor (TNF) can be detrimental in HIV-infected patients, we investigated the effects of melanocortins on production of IL-1 and TNF alpha in the blood of HIV patients. Cytokine production was measured in whole blood samples stimulated with LPS in the presence or absence of alpha-MSH (1-13), alpha-MSH (11-13), ACTH (1-24), or ACTH (1-39). Melanocortins reduced production of both cytokines in a concentration-dependent fashion. In separate experiments on normal peripheral blood mononuclear cells (PBMC), alpha-MSH (1-13) inhibited production of IL-1 beta and TNF alpha induced by HIV envelope glycoprotein gp 120. These results suggest that stimulation of melanocortin receptors in inflammatory cells could be a novel way to reduce production of cytokines that promote HIV replication.

ACTH (1-24) induces cell shape changes in the immunocytes of the bivalve mollusc, Mytilus galloprovincialis. Using computer-assisted microscopic image analysis, we have found that the G protein antagonist suramin sodium, the adenylate cyclase inhibitor 2',5'-dideoxyadenosine, and the protein kinase inhibitor staurosporine inhibit this effect. The highly specific inhibitors H-89 (for protein kinase A) and calphostin C (for protein kinase C) only inhibited partially the morphological alterations. In contrast, the simultaneous action of H-89 and calphostin C completely blocked these changes. The above findings indicate that ACTH (1-24) induces cell shape changes in molluscan immunocytes via adenylate cyclase/cAMP/protein kinase A pathway, as well as the activation of protein kinase C.

In many animal species, the ICV injection of ACTH and of several shorter sequences of the ACTH molecule (melanocortin peptides) induces a peculiar behavioral syndrome mainly characterized by excessive grooming and by repeated acts of stretching and yawning. In adult males, spontaneous penile erections with ejaculation are also induced. We have studied the effect of NO synthase inhibition on this behavioral syndrome. The IP injection of the NO synthase inhibitor L-NG-nitroarginine methyl ester (NAME) significantly prevented--at the doses of 50 and 100 mg/kg--all the behavioral symptoms induced by the ICV administration of ACTH(1-24) (4 micrograms/rat). On the other hand, the ICV injection of NAME (up to 300 micrograms/rat) had no influence on the ACTH-induced excessive grooming and stretching, while significantly inhibited the display of yawnings and penile erections. These data indicate that brain NO synthase is involved in the mechanism of ACTH-induced yawning and penile erections, whereas peripheral NO synthase is involved in the induction of stretching and grooming.

Using ELISAs for B-50/GAP43 and neurofilament (NF), we tested ACTH(1-24), alpha-MSH, ACTH(4-10), and an ACTH(4-9) analogue (ORG2766) for their ability to induce sprouting and neuritogenesis from spinal and sensory neurons. Dissociated fetal rat spinal cord neurons or neonatal rat dorsal root ganglion (DRG) cells were cultured with peptide and assayed after 24, 48, or 96 h. In spinal neurons, alpha-MSH and ACTH(1-24) induced the expression of B-50 dose dependently. After 24 h alpha-MSH had a stimulatory effect (from 10 nM onwards), with a maximum at 100 microM (36% increase). After 96 h the maximal effect of 100 microM alpha-MSH on B-50/GAP43 was lower (19%). ACTH(1-24) (100 microM) stimulated B-50/GAP43 by 19%. Neurofilament levels (96 h) were elevated maximally by 64% at 100 microM alpha-MSH. In DRG neurons a bell-shaped dose-response curve was found for alpha-MSH, the maximal effect being observed after 48 h at 100 nM: 54% for B-50/GAP43 and 22% for NF. In both culture systems neither ACTH(4-10) nor ORG2766 was effective. We conclude that alpha-MSH stimulates the expression of B-50/GAP43 (sprouting) and the formation of NF (neurite elongation) and may therefore be considered a neurotrophic factor.

In view of the increasing evidence that a variety of stresses can influence immune responses, the direct effect of adrenocorticotropic hormone on the migration of human monocytes was studied in vitro. ACTH(1-24) significantly increased the number of migrating cells when placed in the same or the opposite compartment of the chemotaxis chamber, maximum activity being obtained at 10(-14) and 10(-8) M. The results indicate that ACTH(1-24) directly and potently stimulates the migration of human monocytes by means of a chemokinetic effect.

The modulatory action of corticotropin (ACTH) on the specific proteolysis of B-50 to B-60 [B-50(41-226)] was investigated using a previously characterized synaptosomal membrane extract [ASP(57-82)] that was highly enriched in B-50, B-50 kinase activity and a B-50 protease. In common with the previously described inhibitory action on B-50 phosphorylation at Ser41, ACTH(1-24) inhibited B-50 proteolysis in a concentration and structurally dependent manner, while ACTH(1-10) and ACTH(11-24) or a combination of both peptides were ineffective. Structural similarities between segments of ACTH(1-24) and B-50(40-55) may explain the inhibitory action of the peptide on B-50 phosphorylation and proteolysis. In addition, calmodulin, which binds to the N-terminus of the dephosphorylated form of the protein, also protected B-50 from degradation.

The effects of centrally administered ACTH(1-24) and ACTH(4-10) on isolation-induced distress vocalizations (DVs) were assessed in the presence or absence of social cues (mirrored and plain environments). A dose-response analysis indicated that ACTH(1-24) at doses of 0.5 nM and above increased DVs relative to controls when the animals were tested in mirrored or social environments which reduce baseline levels of calling. This effect, however, was short-lived (approx. 15 min). When tested again 1 hr after injection, the treated animals did not differ from controls. ACTH/MSH(4-10) had no effect on vocalization when the animals were tested immediately after injection, but marginally increased calling when animals were tested an hour later. In addition to vocalization changes, ACTH(1-24) induced squatting when animals were isolated in the test boxes, and yawning, head shaking, wing flapping and preening when animals were reunited after testing. ACTH(1-24)-treated chicks also exhibited longer latencies to close their eyes when they were held in the cupped hands of the experimenter. Taken together, the results suggest that ACTH(1-24) induces a central state of arousal in chicks that resembles fear/anxiety.

Experiments aimed at determining the neural basis of reward have previously focused on the role of neurotransmitters and have only recently begun to investigate the role of peptides. The present experiment investigated the effect of ACTH1-24 on d-amphetamine self-administration in rats. Animals were trained daily (8 hour sessions) to press a lever which activated a system that administered 0.125 mg/kg of intravenous amphetamine. After achievement of a stable self-injection frequency, subjects were injected SC with 10, 20 or 40 micrograms/80 microliters ACTH1-24 immediately prior to placement in the apparatus. The 20 micrograms and 40 micrograms doses of the peptide fragment induced a statistically significant attenuation of d-amphetamine self-injection which lasted for 2 days. Control rates of responding were achieved by 5 to 10 days after the peptide treatment. An experiment was conducted to evaluate possible neuromodulatory effects of the peptide fragment. Twenty-four hr after ACTH1-24, HVA was elevated in the caudate. When both apomorphine and ACTH1-24 were administered, the combination lowered HVA in the caudate to a greater degree than apomorphine alone. The peptide fragment, when combined with haloperidol, attenuated the haloperidol-induced increases of DOPAC and HVA in both the caudate and nucleus accumbens. It was tentatively concluded that the neuromodulatory action of ACTH1-24 on dopaminergic neurons may result in an increase in the rewarding quality of d-amphetamine, thus rendering control level self-infusions superfluous.(ABSTRACT TRUNCATED AT 250 WORDS)

ACTH-(1-24) inhibits the in vitro binding of the muscarinic antagonist [3H]QNB to membranes from rat brain. The magnitude of inhibition is dependent on the concentration of ACTH-(1-24). Kinetic analysis indicates a pure competitive inhibition which is suggestive of a reversible interaction of ACTH with muscarinic receptors. A mechanism involving an interaction of ACTH-(1-24) with the phospholipid core of the receptors is suggested. Structure activity studies point to a relation with reported effects of intracerebroventricularly administered ACTH on the turnover rate of acetylcholine and the ACTH-induced stretching and yawning syndrome.

The gross behavior induced by centrally administered bombesin in rats was compared to that elicited by ACTH-(1-24) and the somatostatin analog, des AA1,2,4,5,12,13[D-Trp8]-somatostatin (ODT8-SS). Bombesin (0.001-1 microgram, ICV) caused dose-related excessive scratching which was qualitatively different from that associated with the other two groom-inducing agents. Bombesin-induced grooming was not markedly affected by behaviorally nondepressant doses of haloperidol, morphine, naloxone or neurotensin. Bombesin was active in genetically hypotrichotic (essentially furless) rats; and, again in such animals, even after numbing the area caudal to the shoulders with lidocaine. Tolerance and cross-tolerance studies with bombesin and ODT8-SS indicated that they produce scratching through different mechanisms. Bombesin caused scratching when injected directly into the periaqueductal gray, but not when administered intravenously. Neither hypophysectomy nor adrenalectomy markedly affected bombesin-induced grooming. This behavior appears to be initiated in the central nervous system and is produced independently of the pituitary-adrenal axis.

ACTH (1--24) and alpha-melanotropin (alpha-MSH), peptides previously shown to influence body temperature when administered centrally and to occur naturally in brain regions important to temperature control, were injected intracerebroventricularly (ICV) in rabbits. The peptides in doses of 1.25, 2.5 and 5.0 micrograms produced dose-related hypothermias in a 23 degrees C environment, and greater decreases in body temperature when the experiments were repeated in the cold (10 degrees C), but the largest dose had no effect on temperature in the heat (30 degrees C). These results indicate that the peptides do not reduce the central set-point of temperature control. Rather, they appear to selectively inhibit heat conservation and production responses. Five microgram of ACTH reversed vasoconstriction and inhibited rises in temperature caused by leukocytic pyrogen (LP) given IV and ICV. The same dose of alpha-MSH also reduced fever produced by IV and ICV LP, but the reduction was not as great as after ACTH. Both peptides (5 micrograms) also reduced temperature rises and vasoconstriction caused by ICV PGE2. ACTH reduced d-amphetamine-induced hyperthermia without altering vasoconstriction which suggests that this peptide can reduce temperature rises by inhibiting heat production alone. One of the most important findings was that the peptides are antipyretic in that they reduce fever at doses (0.25 microgram, ICV) that do not affect normal temperature. The powerful effects of these peptides on resting body temperature, hyperthermia and fever, together with their presence in brain tissue important to temperature control, suggest that the endogenous central peptides participate in thermoregulation, perhaps by limiting fever and influencing normal temperature.

We have investigated the possibility that opiate peptides induce excessive grooming behavior in the rat via a direct action on an opiate receptor by comparing the opiate agonist dynorphin(1-13) with its non-opioid fragment des-tyrosine1-dynorphin(1-13) (dT-Dyn). We have shown that both peptides are capable of inducing grooming and that this behavior can be suppressed by pretreatment with naloxone. Analysis of the grooming pattern revealed that the response induced by dT-Dyn is qualitatively similar to that induced by ACTH(1-24) and dynorphin(1-13). Cross-tolerance was demonstrated among the various peptides. We conclude that peptide-opiate receptor interaction is not the primary event in the induction of grooming and that the opiate receptor(s) involved are located at another site underlying peptide-induced grooming.

Bombesin and ACTH-(1-24) induce a dose dependent increase in grooming behavior. Lower doses of bombesin induce a more general type of compulsive grooming in which most elements are involved, whereas higher amounts of bombesin induce a shift towards the element scratching at the cost of bodily grooming and sexual grooming. In contrast ACTH-(1-24) induces a dose dependent increase of all elements of grooming. It is concluded that the grooming displayed by animals treated with ACTH-(1-24) or with bombesin is of a completely different nature. In addition it is observed that under the conditions used tolerance occurs for the grooming inducing effect of ACTH-(1-24), but not for that of bombesin. Moreover, it appears that no cross tolerance exists between bombesin and ACTH-(1-24).

We studied the grooming response to lateral ventricle injection of CRF in both rats and mice under similar conditions. One microgram of CRF ICV induced a pronounced increase (3- to 4-fold) in the frequency of self-grooming in rats, but only a much smaller (less than 20%) increase in mice. The minimum effective dose of CRF in rats was 300 ng. Although ACTH1-24 induced less grooming in mice than in rats, the difference in potency did not appear to be sufficient to explain the differences between the effectiveness of CRF in the two species. Whereas ACTH increased all types of grooming scored. CRF increased all forms of grooming except flank scratching with the hind limb. The major effect of CRF was to increase the number of episodes of grooming, whereas ACTH1-24 tended to prolong the length of individual episodes. The excessive grooming induced by ICV CRF was not affected by prior treatment with dexamethasone, suggesting that the increased grooming was not due to secondary release of ACTH from the pituitary. Nevertheless, ICV CRF might induce grooming by releasing MSH/ACTH from cerebral storage sites. CRF-induced grooming, like ACTH-induced grooming, was inhibited by naloxone pretreatment. Despite the small qualitative differences, CRF-induced grooming could be due to secondary release of ACTH.

Sixteen peptides were injected intracerebroventricularly to test their effects on rectal temperature of rabbits in a thermoneutral environment. In initial tests 5 micrograms alpha-MSH, ACTH(1--24), oxytocin, vasopressin and glucagon altered body temperature while ACTH(1--10), cholecystokinin, contraceptive tetrapeptide, gastrin, insulin, interferon, leupeptin, LHRH, panhibin (somatostatin), and proctolin did not. Bombesin also altered body temperature but in no consistent direction. In further tests on the effective peptides 1.25--5.0 micrograms alpha-MSH and ACTH(1--24) produced dose-related decreases in rectal temperature as great as 1.0 degrees C. The same doses of oxytocin and glucagon produced small, prolonged hyperthermias which did not exceed 0.4 degrees C. Vasopressin caused rapid development of small increases in rectal temperature; temperature returned to normal in 2--3 hr. The results suggest that five of the peptides tested may have roles in central mediation of normal body temperature, hypothermia, hyperthermia and fever.

ACTH-(1-24) and alpha-MSH, intracerebroventricularly (ICV) injected at the doses of 4 and 10 micrograms/animal, respectively, markedly inhibited spontaneous feeding in adult Sprague-Dawley rats, the effect remaining significant for 6-9 hours. At these same doses, ACTH-(1-24) and alpha-MSH abolished the feeding-stimulatory effect of the kappa opiate receptor agonist pentazocine, intraperitoneally (IP) injected at the dose of 10 mg/kg. The same antagonism was obtained by ICV injection of ACTH-(1-24) into rats IP treated with other kappa opiate agonists, bremazocine and tifluadom, at the doses of 1 and 5 mg/kg, respectively. These data suggest that melanocortin peptides play an inhibitory role in the complex regulation of food intake, and further support and extend the hypothesis of a melanocortin-opioid homeostatic system, its two neuropeptide components usually having opposite, mutually-balancing effects.

Experiments were conducted to compare the blood pressure and heart rate responses of conscious rats given intracerebroventricular (ICV) injections of adrenocorticotropin (ACTH 1-24) and corticotropin releasing factor (CRF). Under sodium pentobarbital anaesthesia, rats were implanted with a stainless-steel cannula into the lateral cerebral ventricle and had their right femoral artery and vein cannulated. Upon recovery (24-48 hr later) conscious, unrestrained rats were given ICV injections (total volume 5 microliter by gravity flow) of sterile saline, ACTH (1-24) (0.85 and 1.7 nmoles) or CRF (0.55 and 1.1 nmoles) and blood pressure and heart rate were monitored over the next 2 hr (from the abdominal aorta via the femoral arterial catheter). Both ACTH and CRF caused mean arterial pressure (MAP) to increase, which was paralleled with increases in mean heart rate (MHR). Moreover, these elevations in MAP and MHR were temporally associated with excessive grooming (for ACTH) and locomotor activity (for CRF), which occurred before and lasted as long as MAP and MHR were enhanced. Intravenous (IV) pretreatment whereby naloxone was given 10 min before ICV administration of ACTH (1.7 nmoles) or CRF (1.1 nmoles), showed that naloxone blocked the behavioral, pressor and tachycardic effects of both ACTH and CRF. The results demonstrate that the pressor, tachycardic and locomotor effects evoked in conscious rats by ICV administration of ACTH or CRF are antagonized by naloxone and that their hemodynamic changes may, in part, be mediated by prior behavioral activation.

Norepinephrine (NE)-induced contractile responses were less in aortic strips from SH compared to WKY rats. ACTH 1-24 potentiated NE responses in both SH and WKY aortic strips. This effect was more potent in SH aortic strips. NE-induced contractions in SH aortic strips were less sensitive to changes in external Ca2+ levels than were those of WKY aortic strips. ACTH 1-24 did not potentiate NE responses under low external Ca2+ conditions in SH aortic strips or under high external Ca2+ conditions in WKY aortic strips. The greater sensitivity of NE responses following ACTH 1-24 in SH aortic strips may imply that this peptide is modulating a mechanism related to an impaired contractility and that Ca2+ plays a key role in the observed effects.