Comparison of the effects of adrenocorticotropic hormone fragment 4-10 (ACTH(4-10)) and [D-Phe7]ACTH(4-10) on the compensation of spontaneous nystagmus in guinea pig

Following unilateral labyrinthectomy (UL), spontaneous nystagmus (SN) was measured in guinea pigs which received i.m. injections of 100 or 400 μg/kg ACTH(4-10) or 800 μg/kg [D-Phe7]ACTH(4-10), every 4 h for 48 h post-UL. The results were compared to those from a previous study, conducted under identical conditions, in which guinea pigs received similar injections of 200 μg/kg ACTH(4-10) or 0.1 ml/kg saline. ACTH(4-10) significantly accelerated the rate of compensation of SN at 100, 200, and 400 μg/kg doses, although the average effects were larger for the 200 μg/kg condition. [D-Phe7]ACTH(4-10) significantly increased the frequency of SN following UL compared to saline-treated animals; however, it also accelerated the rate of SN compensation. These results indicate that the acceleration of SN compensation in guinea pig by ACTH(4-10) follows an inverted U-shaped dose-response curve and that [D-Phe7]ACTH(4-10) increases the frequency of SN.     

Effect of ACTH-like peptides on morphine-induced hypothermia in unrestrained guinea pigs

Peripheral treatment with adrenocorticotropin (1-24) (ACTH1-24), at different doses and sequences, consistently antagonized the decrease in body temperature produced by morphine in the freely moving guinea pig, whereas adrenocorticotropin (4-10) (ACTH4-10), which lacks corticotrophic activity, was partially effective only when it was administered in a high dose 24 h prior to morphine. Centrally administered ACTH1-24 completely prevented the hypothermic effect of intracerebroventricularly (i.c.v.)-injected morphine. Likewise, the i.c.v. administration of ACTH4-10 was equally effective in blocking the i.c.v. morphine-induced hypothermia. Neither ACTH1-24 nor ACTH4-10 did produce changes in body temperature. These results suggest that peripherally administered ACTH1-24 antagonizes indirectly the actions of morphine through the release of adrenal corticosteroids, whereas centrally injected ACTH1-24 or ACTH4-10 act as direct antagonists of morphine effects through opioid receptors.     

Different effects of ACTH fragments on hippocampal EEG and behavior

The hippocampal electrical activity and behavior of rabbits have been studied in the presence of novel and emotional stimuli. The effects of these stimuli have been recorded in controls and in groups of animals treated with ACTH (4-10)(10 micrograms/100 g, i.m.) and ACTH (1-24) (10 micrograms/100 g, i.m.). Recordings were made immediately and 30 min after injection. ACTH (4-10) injection failed to evoke any significant behavioral or electrical response. Rabbits recorded 30 min after ACTH (1-24) injection showed a reduction of hippocampal RSA (rhythmic slow activity) frequency and behavioral activity. In particular, reduction of exploration, self-grooming, motor activity and approaches to the new object have been observed. Since ACTH is characterized by a corticotropic action these results can be due to corticosteroid stimulation.     

Character of (D-Phe-7) ACTH4-10-induced excessive grooming

Rats receiving (D-Phe-7) ACTH4-10 exhibited excessive grooming during the first half of the hour-long observation period. This resulted in total grooming scores of about one-half of those produced by the longer ACTH fragments, ACTH1-24 or ACTH1-16 NH2. The excessive grooming induced by (D-Phe-7) ACTH4-10 was due to an enhancement of duration of grooming bouts and not to an increase in the frequency of occurrence. Furthermore, the neuropeptide-induced excessive grooming was eliminated by prior treatment with naloxone.     

ACTH-like neurotropic peptides: possible regulators of rat brain cyclic AMP

The influence of behaviorally active, N-terminal fragments of ACTH on the accumulation of cAMP in rat brain investigated in broken cell preparations of subcortical tissue, in slices of neostriatum and in vivo. ACTH1--24 has a biphasic effect on the activity of adenylate cyclase in broken cell preparations of rat brain subcortical tissue: concentrations below 25 micrometer stimulated, whereas concentrations of 0.1 mM and higher inhibited adenylate cyclase activity. The magnitude of the stimulation was dependent on the concentrations of ATP and Mg2+ in the incubation medium. Structure activity studies revealed that at a concentration of 10(-4) M ACTH1--16-NH2 and ACTH4--7 also inhibited the activity of adenylate cyclase, whereas ACTH11--24, ACTH1--10, ACTH4--10, [D-Phe7]ACTH1--10 and [D-Phe7]ACTH4--10 were inactive in this respect. Addition of 0.8 mM EGTA but not of 0.25 mM Ca2+ prevented the inhibition by 10(-4) M ACTH1--24. GMP-N-P (10(-5) M), naltrexone (10(-3) M) and ergometrine (10(-3) M) did not influence the inhibitory effect. ACTH1--24 enhanced the accumulation of cAMP in slices from rat brain neostriatum in a dose-dependent manner. This effect was already maximal 7.5 min after the addition of the peptide and was potentiated by isobutylmethylxanthine, a potent inhibitor or phosphodiesterase. Intraventricular injection of 1 microgram ACTH1--16-NH2 in rats significantly elevated (+ 27%) the concentration of cAMP in the septal region 60 min after the injection of the peptide. The results are discussed in terms of a possible involvement of cAMP as a second messenger in the central nervous system for N-terminal fragments of ACTH.     

Cerebral 2-deoxyglucose accumulation in mice following chronic treatment with an ACTH4–9 analog, ORG 2766

Mice were treated with [MET(O2)4, D-Lys8,Phe9]ACTH4-9 (ORG 2766, 100 micrograms/kg per day SC) for 10 days. On the tenth day mice were injected with [3H]2-deoxyglucose and its cerebral accumulation determined in 17 brain areas. The combined results of four experiments indicated a significant decrease of the 2-deoxyglucose accumulation in the septum. Changes observed in other brain areas were not statistically significant in the combined analysis of all four experiments. This selective change in the septum is consistent with selective uptake of ORG 2766 in this region, and with the lack of behavioral activity of ACTH in septal lesioned animals. It also suggests that the behavioral activity of this peptide, generally considered to be on arousal, vigilance, and/or selective attention, may be mediated through the septum, a limbic system structure.     

Social and exploratory behaviour in the rat after septal administration of ORG 2766 and ACTH4-10

The time spent in active social interaction by pairs of male rats in a dimly-lit, familiar test arena, was decreased significantly after intraseptal injection of ACTH4-10 (250-500 ng). In contrast, the time spent in active social interaction was increased significantly after intraseptal injection of the tri-substituted ACTH4-9 analogue ORG 2766 (250-500 pg). Neither ACTH4-10 (250 ng) nor ORG 2766 (250 pg) affected exploration or locomotor activity measured in a holeboard, after intraseptal injection. Intraseptally administered ACTH4-10 (250 ng) increased aggression measured in a colony intruder model, but ORG 2766 (250 pg) was without effect.     

Effects of neonatal ACTH4-10 administration on adult adaptive behavior and brain tyrosine hydroxylase activity

The effects of daily injection of ACTH4-10 in postnatal days 2-8 were studied on adult adaptive behavior and brain TH activity in male rats. Four groups were used: (1,2) 10 and 2 nmol ACTH4-10 injected, (3) saline injected, and (4) intact, unhandled. The peptide did not influence acquisition but delayed extinction of CAR in the shuttle-box and facilitated passive avoidance behavior against saline control. The experimental manipulation in the saline control group (neonatal stress) also significantly influenced these behavioral variables and motility during conditioning but in the opposite direction as did ACTH4-10 treatment. It was concluded that ACTH4-10 interfered with the action of neonatal stress on the development of adaptive behavior. A similar kind of interaction could also be observed in TH activity of striatum and locus ceruleus but not in all brain areas investigated.     

Interactions of ACTH4-10 and ACTH1-24 with L-[3H]glutamate binding sites and GABA/benzodiazepine/picrotoxin receptor complexes in vitro

The effects of ACTH4-10 and ACTH1-24 on L-[3H]glutamate (Glu) binding sites and GABA/benzodiazepine/picrotoxin receptor complexes in vitro were investigated. ACTH4-10 and ACTH1-24 inhibited [3H] Glu and [3H] muscimol binding concentration-dependently, while [3H] flunitrazepam (FNP) and [35S] t-butylbicyclophosphorothionate (TBPS) binding were not affected. These ACTH fragments also inhibited GABA-stimulated [3H] FNP binding. These results suggest that ACTH and its fragments may act as anticonvulsants by antagonizing glutamate binding, their interaction with GABA-A sites may relate to the other central nervous effects of ACTH than the anticonvulsant activity.     

ACTH and neuromuscular transmission: electrophysiological in vitro investigation of the effects of corticotropin and an ACTH fragment on neuromuscular transmission.

The effects of corticotropin (ACTH) and the polypeptide fragment ACTH 4-10 on neuromuscular transmission were studied in rat phrenic nerve–diaphragm preparations in vitro. ACTH decreased the quantum content of end-plate potentials (epps) and increased the transmission failure rate. The frequency of miniature end-plate potentials (mepps) was increased. The results indicate that ACTH acts directly on the presynaptic terminal. The findings could explain the transient decrease in muscle strength often observed during corticotropin therapy for myasthenia gravis, and they suggest that the therapeutic effect of ACTH is indirectly mediated. The only marked effect produced by the ACTH fragment was an increase in the mepp frequency; quantum content and failure rate of epps remained unchanged. Since ACTH 4-10 is unable to stimulate cortisone release, its clinical efficacy seems unlikely.     

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.     

EEG and task performance after ACTH4-10 in man.

Effects of the heptapeptide ACTH4-10 (Org OI-63) on EEG, some memory tests, and behavior were examined in 12 normal male volunteers under 30 years of age. An intravenous dose of 60 mg was compared to placebo in a Latin square design. EEG was recorded for 2 h after administration of ACTH4-10 or placebo and was quantified by power spectral density analysis. Drug differences were tested by analyses of variance and covariance. No statistical drug effect was seen on the EEG or behavioral measures. Of the psychological tests, only the digit span test showed a decrease in number of errors with ACTH4-10 (p less than or equal to o.05). These results are consistent with previous studies and suggest that an intravenous bolus of ACTH4-10 has a limited effect on the brain function tested.     

Lipolytic potency of proopiomelanocorticotropin peptides in vitro

Four lipolytic active centers were localized in proopiomelanocorticotropin (POMC): in the N-terminal part of POMC ("tryptophane rich peptide"), in the N-terminal part of adrenocorticotropic hormone, in the middle portion of beta-lipotropin and in the C-terminal part of beta-lipotropin. The weak lipolytic activity of the "tryptophane rich peptide" is not mediated by its two partial sequences gamma-MSH and delta-MSH. The minimal amino acid sequence for obtaining lipolysis from the N-terminal part of ACTH was ACTH 4-10. Lengthening of this amino acid sequence on the N- or C- terminus resulted in a strong increase of lipolytic potency. The minimal effective sequence from the middle portion of beta-lipotropin was located in the amino acid residues 47-53 which are identical to ACTH 4-10. Additional amino acids on the N- and C-terminus (beta-lipotropin p 41-58 and beta-lipotropin h 35-56) lead also to increased lipolytic activity. The forth center of POMC resides in the C-terminal part of beta-lipotropin (residues 78-91) because sequences from the N-terminal part of beta-lipotropin 61-91 were ineffective. The order of potency of POMC peptides especially in respect to the minimal effective concentration was ACTH 1-13 (alpha-MSH) greater than beta-lipotropin p greater than ACTH greater than ACTH 1-10 greater than beta-lipotropin p 61-91.     

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.     

ACTH4-10 and memory in ECT-treated and untreated patients. I. Effect on consolidation

The aim of the study was to investigate the possible effect of a single dose of ACTH4-10 (30 mg subcutaneously) on memory as measured with a test battery appropriate for the study of "consolidation". Twenty depressive inpatients participated in the intraindividual double-blind cross-over comparison between ACTH4-10 and placebo in connection with the second and third treatment in the ECT series. ACTH4-10 (or placebo) was administered subcutaneously 30 min after unilateral ECT. Memory functions were examined 90 min before and 150 min after the administration of the drug. A similar design was used in a study of 20 inpatients with neurotic depression not treated with ECT. The results of both studies give no evidence of a positive influence of a single dose of ACTH4-10 on consolidation of memory. Possible sources of error are discussed.     

Specific uptake of a behaviorally potent [3H]ACTH4-9 analog in the septal area after intraventricular injection in rats.

Distribution within the brain of a behaviorally potent [3H]ACTH4-9 analog 2 h after intraventricular injection in rats was studied in the presence and absence of behaviorally and structurally similar peptides, to explore the significance of earlier found preferential uptake of the [3H]ACTH4-9 analog in the septal area. Hypophysectomy resulted in significantly enhanced uptake of radioactivity in the septum as compared to normal rats. No increase in this brain area of hypophysectomized rats was observed after intraventricular injection of [3H]Phe. Elevated circulating ACTH levels after adrenalectomy seemed too low to compete with the septal uptake of the ACTH4-9 analong. Subcutaneous substitution of hypophysectomized rats with sustained release zinc phosphate preparations of the behaviorally equipotent peptides ACTH1-24 and ACTH4-10 decreased the accumulation of the [3H]ACTH4-9 analog in the septum, whereas treatment with the behaviorally inactive fragment ACTH11-24 is not effective. Retreatment of hypophysectomized rats with neuropeptides, differing structurally from natural ACTH peptides (7-D-Phe-ACTH4-10, BETA-LPH61-76 and 9-desglycinamide, 8-Lys-vasopressin), did not change the uptake of the ACTH4-9 analog in any of the investigated brain areas. These results give evidence for specific uptake of the ACTH4-9 analog in the septal region, because competitive displacement occurs only with peptides which both behaviorally and structurally are closely related to the ACTH4-9 analog.     

The effect of ACTH4–10 on protein synthesis, actin and tubulin during regeneration

The effects of ACTH4-10, a peptide fragment of corticotropin, on rat dorsal root ganglia (DRG), spinal cord and sciatic nerve were studied following a crush lesion of the sciatic nerve. The in vitro total protein synthesis rate of DRG L4, L5 and L6, measured one and three days after ipsilateral nerve crush, were not altered by various ACTH4-10 treatment regimes. Likewise, neither ACTH4-10 treatment of sham-operated rats nor in vitro exposure of control ganglia to peptide, resulted in changes in synthesis rate. Four days after crush lesion, the amounts of actin and tubulin in the ventral horn L2-L5 region of the spinal cord and of actin in DRG L5 were estimated following 2-dimensional separation. No significant effect of ACTH treatment was found. Degeneration-associated changes in the protein profiles of segments of sciatic nerve were not altered by ACTH4-10 treatment. The data are discussed in relation to the possible site of action of neurotrophic ACTH-like peptides.     

Alpha-melanocyte-stimulating hormone and behavior

Melanocyte-stimulating hormone (MSH) has putative adaptive significance in all forms of species where it is present. In mammals the polypeptide chain influences learning, memory and attention. Chemically MSH shares the first 13 (alpha-MSH) or the first 18 or 22 (beta-MSH) amino acids with adrenocorticotropic hormone (ACTH), even though the mechanisms of secretion and behavioral effects are often quite different. The still shorter peptide chain MSH/ACH4-10 demonstrates significant melanotropic and behavioral actions of alpha-MSH without showing any steroidogenic effect. Behaviorally, alpha-MSH and MSH/ACTH analogues (MSH/ACTH4-9 and MSH/ACTH4-10 influence the parameters of learning, attention and memory in both human and infrahuman subjects. Alpha-MSH has also been reported to increase sensitivity and augment arousal mechanisms in the CNS. Alpha-MSH has been observed to increase and sustain novelty-induced defecation, and this behavior was found to be accompanied by a concomitant decrease in whole brain DA and NE levels in both intact and hypophysectomized rats exposed daily to a test box. The behavioral effects of alpha-MSH may be partially modulated by the enhanced cyclic-AMP activity in the CNS observed after MSH administration. MSH also seems to be working in conjunction with the hypothalamic tripeptide MIF-1 and the pineal hormone melatonin, both of which can affect the release of MSH from the pituitary. Recent evidence suggests that MSH is counterbalancing against and complementing with the effects of endorphins, specifically beta-endorphin (61-91 chain of beta-LPH), in maintaining learning and attentive behaviors.     

Effect of Antiepileptic and Antimyoclonic Drugs on Serotonin Receptors in vitro

To determine if the mechanism of action of clinical and investigational antiepileptic and antimyoclonic drugs or neuropeptides involves direct actions at serotonin (5-HT) receptors, the activity of various compounds in vitro at 5-HT1 (with subtypes) and 5-HT2 sites was measured in adult rat brainstem, spinal cord, and neocortex. Adrenocorticotropic hormone (ACTH1-39) noncompetitively inhibited specific binding at 5-HT1, 5-HT1A, and 5-HT2 sites in brainstem and neocortex [concentrations required to displace 50% of ligand binding (IC50S) 4-8 X 10(-5) M]. ACTH1-24, ACTH1-17, and ACTH4-10 were sequentially less active, and ACTH34-39 and corticosterone were inactive. D-Ala2, Leu5-enkephalinamide, but not D-Ala2, Met5-enkephalinamide, also displaced spinal and neocortical 5-HT2 sites (IC50 6 X 10(-5) M). Piracetam, glycine, and the clinical antiepileptics valproate, phenacemide, phenytoin, carbamazepine, phenobarbital, diazepam, clonazepam, nitrazepam, and ethosuximide did not displace serotonergic radioligands, but melacimide showed some activity at 5-HT1 sites (IC50 7-9 X 10(-5) M). Anticonvulsant inactivity at 5-HT receptors in vitro correlates with the lack of antimyoclonic activity in 5-HT lesion myoclonic models but not with antimyoclonic efficacy in humans. These data indicate that acute effects of these anticonvulsants cannot be attributed to direct action at the 5-HT receptor recognition site in the rat. In contrast, ACTH showed mild in vitro displacement and regional specificity but only at micromolar concentrations.     

Modulation of the immune response by proopiomelanocortin derived peptides. II. Influence of adrenocorticotropic hormone on the rise in intracellular free calcium concentration after T cell activation

In this paper the effect of ACTH 1-24 is described on one of the "early events" occurring during T cell activation: the rise in the concentration of cytosolic free calcium. The increase in cytosolic free calcium induced by concanavalin A or by a monoclonal antibody with specificity for the T cell receptor-T3 complex could be modulated by physiological concentrations of ACTH 1-24. The modulation of the response appeared to be dose dependent. The effect of ACTH 1-24 was maximal at 10(-10) M. No effect of ACTH 1-24 was observed on the basal level of cytosolic free calcium. These results indicate that physiological doses of ACTH can interfere with the responsiveness of T cells to mitogenic and antigenic stimuli.