Brain Opioid Peptides Research Articles

Mammalian neurotoxins, Blarina paralytic peptides, cause hyperpolarization of human T-type Ca channel hCav3.2 activation

Among the rare venomous mammals, the short-tailed shrew Blarina brevicauda has been suggested to produce potent neurotoxins in its saliva to effectively capture prey. Several kallikrein-like lethal proteases have been identified, but the active substances of B.brevicauda remained unclear. Here, we report Blarina paralytic peptides (BPPs) 1 and 2 isolated from its submaxillary glands. Synthetic BPP2 showed mealworm paralysis and a hyperpolarization shift (-11mV) of a human T-type Ca2+ channel (hCav3.2) activation. The amino acid sequences of BPPs were similar to those of synenkephalins, which are precursors of brain opioid peptide hormones that are highly conserved among mammals. However, BPPs rather resembled centipede neurotoxic peptides SLPTXs in terms of disulfide bond connectivity and stereostructure. Our results suggested that the neurotoxin BPPs were the result of convergent evolution as homologs of nontoxic endogenous peptides that are widely conserved in mammals. This finding is of great interest from the viewpoint of the chemical evolution of vertebrate venoms.

Treatment with steroid hormones and morphine alters general activity, sexual behavior, and opioid gene expression in female rats

AimsPrevious studies have shown that brain opioid peptides exert an inhibitory influence on gonadotropin secretion. Different types of brain opioids, such as β-endorphin, enkephalin, and dynorphin, exert their actions by binding to specific opioid receptors (i.e., μ, δ, and κ, respectively). The present study determined the effects of chronic treatment with morphine in female rats with pharmacologically induced estrus on behavior and opioid receptor gene and protein expression in the hypothalamus, striatum, and periaqueductal gray. Main methodsFemale ovariectomized rats treated with estrogen+progesterone received 3.5mg/kg morphine once per day for 6days. We evaluated general activity, sexual behavior, Oprm1, Oprd1, and Oprk1 gene expression, and μ opioid receptor (MOR), δ opioid receptor (DOR), and κ opioid receptor (KOR) protein expression in the hypothalamus, striatum, and periaqueductal gray in adult virgin female ovariectomized rats. Key findingsChronic morphine treatment increased locomotion and grooming behavior, decreased immobility time, decreased sexual behavior, and decreased the lordosis quotient. The molecular biology results showed that morphine treatment increased Oprm1 gene and MOR protein expression in the striatum and decreased KOR protein expression in the hypothalamus in animals that were assessed for general activity. The animals that were evaluated for sexual behavior exhibited an increase in Oprm1 expression in the periaqueductal gray and increase in KOR expression in the striatum. SignificanceThese results suggest that both opioid system activation and sex hormones alter behavioral and molecular patterns in ovariectomized rats within a relatively short period of time.

Precursors of molecules related to mammalian opioid peptides in brain of a marine worm.

Total mRNA were extracted from brain of Nereis diversicolor (Annelida, Polychaeta) and were translated in vitro or in ovo. The newly synthesized polypeptides were analyzed through electrophoresis of immunoprecipitated products or the Western blotting technique using polyclonal antibodies raised against mammalian dynorphin 1-17 and mammalian alpha-neo-endorphin. Among the products translated in vitro, only one class of polypeptide of 70 kDa was recognized by anti-dynorphin 1-17 antibodies. Furthermore, some in ovo translated products as well as proteins extracted from brain of worms showed identical immunoreactivity. These polypeptides, 60-70 kDa, reacted with anti-dynorphin 1-17 and anti alpha-neo-endorphin antibodies. These results suggest the existence of epitopes common to in ovo and in vitro translated products, to polypeptides extracted from the brain and to some mammalian opioid peptides of the prodynorphin family. We postulate the presence, in the brain of N. diversicolor, of precursors of peptides related to mammalian dynorphin 1-17 and alpha-neo-endorphin. Data reported in this investigation do not allow us to propose or even postulate the presence, in the brain of the worm, of one precursor molecule common to polypeptides related to mammalian dynorphin 1-17 and alpha-neo-endorphin. Furthermore, the Nereis precursor molecules exhibit a clear-cut difference in molecular mass with the mammalian prodynorphin: 70 kDa versus 30 kDa.

Mesolimbic dopamine drives the diurnal variation in opiate-induced feeding

Brain opioid peptides modulate feeding behavior and opiate drugs have powerful orexigenic effects in mammals. Recent studies have shown that opiate-induced eating depends, though not exclusively, on mu-opioid receptors located in the ventral striatum. Here we report that morphine orexigenic effects vary with the time of day according to a biphasic pattern. The effects first increase and then decrease during the light phase, with the peak effect occurring in the middle of this phase. This diurnal profile is shifted toward the dark phase after dopamine deafferentation of the ventral striatum. Consequently, the peak effect of morphine is delayed and occurs just before the dark phase. This finding suggests that mesolimbic dopamine transmission contributes to the neural mechanisms that normally drive the circadian timing of opioid-dependent feeding.

Maternal separation alters maternal care, but has minor effects on behavior and brain opioid peptides in adult offspring

The aim of the study was to investigate the effect of repeated maternal separation (MS; 4 hr per day) during postnatal Days 1 to 15 on emotionality and voluntary ethanol intake in adult male and female Wistar rat offspring relative to controls exposed to a brief (5-min) daily handling procedure. Brain immunoreactive opioid peptide levels and plasma levels of corticosterone also were measured. There were mainly no alterations in any of the tested behaviors (i.e., fleeing and freezing responses, exploratory behavior, spontaneous and amphetamine-induced locomotor activity and competitive behavior), ethanol intake, or immunoreactive opioid peptide levels in MS offspring, either in males or females, compared to their respective controls nor were there any differences in plasma corticosterone between groups. In addition, the dams' retrieval behavior of the pups also was studied, showing that MS dams spent more time in the nest with the pups after the 4-hr separation period compared to control dams. With respect to the used protocol of the MS procedure in the present study, our results do not provide support for the suggestion that this procedure is a relevant model for studying development of psychopathology and vulnerability to drug abuse.

Restricted daily consumption of a highly palatable food (chocolate Ensure(R)) alters striatal enkephalin gene expression.

Brain opioid peptide systems are known to play an important role in motivation, emotion, attachment behaviour, the response to stress and pain, and the control of food intake. Opioid peptides within the ventral striatum are thought to play a key role in the latter function, regulating the affective response to highly palatable, energy-dense foods such as those containing fat and sugar. It has been shown previously that stimulation of mu opiate receptors within the ventral striatum increases intake of palatable food. In the present study, we examined enkephalin peptide gene expression within the striatum in rats that had been given restricted daily access to an energy-dense, palatable liquid food, chocolate Ensure(R). Rats maintained on an ad libitum diet of rat chow and water were given 3-h access to Ensure(R) daily for two weeks. One day following the end of this period, preproenkephalin gene expression was measured with quantitative in situ hybridization. Compared with control animals, rats that had been exposed to Ensure(R) had significantly reduced enkephalin gene expression in several striatal regions including the ventral striatum (nucleus accumbens), a finding that was confirmed in a different group with Northern blot analysis. Rats fed this regimen of Ensure(R) did not differ in weight from controls. In contrast to chronic Ensure(R), acute ingestion of Ensure(R) did not appear to affect enkephalin peptide gene expression. These results suggest that repeated consumption of a highly rewarding, energy-dense food induces neuroadaptations in cognitive-motivational circuits.

Long-term effects of short and long periods of maternal separation on brain opioid peptide levels in male Wistar rats

Environmental manipulations early in life may induce persistent alterations in adult behaviour and physiology. The underlying neural mechanisms of these responses are not yet clear. We have previously reported long-term changes in brain opioid peptide levels in male and female Sprague–Dawley rats after short periods (15 min, known as neonatal handling) of maternal separation (MS) until weaning. To study this further, we investigated behavioural and neurochemical effects of repeated MS in male Wistar rats. The rat pups were separated from their dams in litters for either 360 min (MS360) or 15 min (MS15) daily from postnatal day 1 to 21 or exposed to normal animal facility rearing. Behavioural analysis showed that MS360 rats had increased ultrasonic calls on postnatal day 5 compared to MS15 rats, but not on postnatal day 6. Moreover, the MS360 rats had more animals with higher frequency of calls at day 5 than 6 than the MS15 rats. Analysis of the opioid peptides dynorphin B and Met-enkephalin-Arg 6Phe 7 with radioimmunoassay 7 weeks after the MS procedure, revealed long-term neurochemical changes in several brain areas and in the pituitary gland. Immunoreactive dynorphin B and Met-enkephalin-Arg 6Phe 7 levels were affected in the hypothalamus and dynorphin B levels in the neurointermediate pituitary lobe, amygdala, substantia nigra and the periaqueductal gray. Together, these findings show that repeated periods of MS early in life in male Wistar rats affect the development of the ultrasonic call response and induce long-lasting and possibly permanent alterations in the opioid peptide systems.

Maternal Separation Alters Acquisition of Ethanol Intake in Male Ethanol‐Preferring AA Rats

Background: Prolonged daily maternal separation can increase the risk for developing substance abuse, whereas brief maternal separation has been reported to induce positive behavioral effects, decrease voluntary ethanol intake and induce long‐lasting changes in brain opioid peptides. The ethanol‐preferring AA (Alko, Alcohol) rats have altered basal levels of endogenous opioid peptides that may relate to their high voluntary ethanol intake. The purpose of this study was to investigate whether maternal separation could affect acquisition of ethanol intake in AA rats.Methods: The rat pups were exposed to 15 min (MS15) or 360 min (MS360) of maternal separation during postnatal day 1‐21, while control rats were exposed to normal animal facility rearing. As adults, the male rats were gradually introduced to increasing concentrations of ethanol. Furthermore, the effect of restraint stress on voluntary ethanol intake was investigated.Results: The MS15 rats reached a high voluntary ethanol intake later than MS360 and control rats. The MS15 rats had a lower ethanol intake and preference at 8% ethanol compared to MS360 rats and lower ethanol intake compared to control rats. MS15 rats also had a lower 10% ethanol intake in comparison with MS360 rats. Restraint stress decreased the ethanol intake in MS15 and MS360 rats, whereas the ethanol intake in control rats was unaffected.Conclusions: We have previously shown that prolonged periods of maternal separation in Wistar rats result in an increased ethanol intake later in life. This was not repeated in this study, using AA rats with an inherent high ethanol intake. However, it is shown that brief maternal separation can delay acquisition of high ethanol intake and in addition decrease voluntary ethanol intake and preference in AA rats. Maternal separation for 15 min is therefore suggested to protect against high voluntary ethanol intake later in life.

Anabolic androgenic steroids affects alcohol intake, defensive behaviors and brain opioid peptides in the rat

The present study investigated whether a relationship exists between nandrolone decanoate and voluntary ethanol intake in laboratory rats. Animals were subjected to daily subcutaneous injections with nandrolone decanoate (15 mg/kg) during 2 weeks. One group of animals was tested for voluntary alcohol intake 1 week after the end of the 2-week treatment period and another group received alcohol 3 weeks after the treatment. In addition, assessment of defensive behaviors and immunoreactivity (ir) levels of the brain opioid peptides dynorphin B and Met-enkephalin-Arg-Phe (MEAP) were performed. The nandrolone decanoate-treated animals were significantly more aggressive and showed lower fleeing and freeezing reaction than the oil-treated controls. Treatment with nandrolone decanoate enhanced voluntary alcohol intake, regardless if it was presented 1 or 3 weeks after end of the treatment period. These animals had a decreased activity of dynorphin B-ir in the nucleus accumbens, decreased levels of MEAP-ir in the periaqueductal gray (PAG) and higher levels of MEAP-ir in the hypothalamus compared to controls. In line with previous studies, this suggests that the altered dynorphin B-ir activity may promote the rewarding effects of ethanol and thereby increasing alcohol intake, whereas MEAP-ir may be associated with the ability to control the aggressive reaction. Abuse of nandrolone decanoate may thus constitute a risk factor for increased alcohol consumption and defensive aggression. In human, this constellation of behavioral symptoms is closely related to acts of crimes and violence and is often observed among those abusing anabolic androgenic steroids.

Brain dynorphin and enkephalin systems in Fischer and Lewis rats: effects of morphine tolerance and withdrawal

Lewis rats are more likely to self-administer various drugs of abuse than Fischer rats. Here these two strains of rats were compared with regard to basal brain opioid peptide levels and the response to chronic morphine treatment and to naloxone-precipitated withdrawal. Lewis rats had lower basal dynorphin peptides in the substantia nogra, striatum (not Leu-enkephalinArg 6) and VTA (not dynorphin B) and the pituitary gland. Leu-enkephalinArg 6 levels were also lower in these structures (with the exception of striatum which had higher levels) and in the nucleus accumbens. There were also strain differences in the response to chronic morphine treatment; in the nucleus accumbens, morphine treatment increased dynorphin A levels in Fischer rats only, in the ventral tegmental area effects were opposite with increased dynorphin levels in Fischer and decreased levels in Lewis rats, in the hippocampus dynorphin levels were markedly reduced in Lewis rats only. In Fischer rats, chronic morphine strongly affected peptide levels in the substantia nigra and striatum, whereas Lewis rats responded less in these areas. Leu-enkephalin, which derives from both prodynorphin and proenkephalin, and Met-enkephalin, which derives from proenkephalin, were effected by chronic morphine mainly in Fischer rats, increasing levels in most of the brain areas examined. The results in this study show (1) strain differences in basal levels of prodynorphin-derived opioid peptides, (2) the prodynorphin system to be differently influenced by morphine in Lewis rats than in Fischer rats and 3) the proenkephalin system to be influenced by chronic morphine in brain areas related to reward processes only in Fischer rats.

Endogenous opioid peptides in brain and pituitary of rats with absence epilepsy

The level of opioid peptides in several brain areas and in the pituitary was estimated in WAG Rij rats, which are considered to be a genetic animal model for human absence epilepsy. In comparison with three groups of non-epileptic controls, these epileptic rats had an elevated level of the proenkephalin-derived peptide Met-enkephalin-Arg 6-Gly 7-Leu 8 in the mesencephalon and striatum, while the level of the prodynorphin-derived peptide α-neoendorphin was increased in the striatum and hippocampus. In addition various age- and/or strain-related changes in these peptide levels were found in the hippocampus, thalamus, striatum, frontal cortex and neurointermediate lobe of the pituitary. No difference in the hypothalamic β-endorphin level were found between epileptic and non-epileptic rats, though strain- and/or age-related changes in the peptide content were detected in both lobes of the pituitary. The increased level of proenkephalin and prodynorphin opioid peptides in brain structures, essential for the appearance of spike-wave discharges, suggests that these opioid systems, but not proopiomelanocortin one, may play a role in absence epilepsy.

Activation of glucocorticold receptors and the effect of naloxone during hemorrhagic hypotension

Evidence indicates that endogenous opioid pcptides and glucocorticoids participate in the control of cardiovascular regulation during hemorrhagic shock. In the present study, we investigated a possible interaction between brain opioid peptides and adrenal corticosteroids regarding the control of arterial pressure during hemorrhage. The bleeding volumes required to lower arterial pressure to 80, 60 and 40 mmHg were studied in anesthetized sham-operated (SHAM) and adrenalectomized (ADX) rats. I.c.v. administration of 10 μg of naloxone resulted in a significant increase in the bleeding volume required to lower arterial pressure from 60 to 40 mmHg in SHAM animals, whereas no effect of naloxone was observed in ADX animals. Replacement therapy with a 100% corticosterone pellet (100 mg, s.c.), but not with a 12.5% corticosterone pellet (12.5 mg corticosterone and 87.5 mg cholesterol, s.c.), resulted in an effect of naloxone on the bleeding volume in ADX animals. The effect of replacement therapy could be inhibited by i.c.v. pretreatment with the synthetic glucocorticoid receptor antagonist, RU38486 (100 ng). These data suggest that (1) opioid mechanisms are involved in the regulation of blood pressure during hemorrhage, and (2) occupancy of glucocorticoid receptors is required for naloxone to exert its hemodynamic effect during hemorrhagic hypotension in ADX rats.

Involvement of brain opioid peptides in blood pressure regulation during controlled haemorrhagic hypotension

Involvement of brain opioid peptides in blood pressure regulation during controlled haemorrhagic hypotension

Positive effects of captopril in the behavioral despair swim test

Captopril, an angiotensin II converting enzyme (ACE) inhibitor, was evaluated for potential antidepressive activity on the forced swim-induced behavioral despair (immobility) test in mice. Captopril (10.0 and 30.0, mglkg ip) significantly reduced immobility and mimicked the effects of the antidepressants imipramine (30.0 mglkg, ip) and mianserin (3.0, 10.0, and 30.0 mglkg, ip). Captopril increased the motor activity of mice at these same dosages. Naloxone (2.0 mglkg, ip) blocked the effects of captopril (30.0 mglkg, ip) in the swim test. These data suggest that captopril has potential antidepressive activity. However, the conclusion is guarded, as the positive effects may be related to motor stimulation. The blockade of the captopril effects by naloxone suggests that brain opioid peptides play a role in this behavioral effect of captopril.

Possible involvement of brain opioid peptides in clonidine-induced hypotension in spontaneously hypertensive rats.

The effect of intracisternal pretreatment with opiate antagonists or antisera against various opioid peptides on the hypotension and bradycardia induced by cumulative intracisternal administration of clonidine (0.02-2.5 microgram) was studied in conscious Wistar-Kyoto rats and in spontaneously hypertensive rats. No effect of any pretreatment on basal values of blood pressure and heart rate was detected in either of these strains. In spontaneously hypertensive rats intracisternal pretreatment with naltrexone resulted in a dose-dependent inhibition of clonidine-induced hypotension and bradycardia. DL-naloxone also antagonized the hypotension but not influence the hypotensive and bradycardic response to clonidine. In Wistar-Kyoto rats naltrexone and the beta-endorphin antiserum B4 failed to affect the cardiovascular effects of clonidine. B4 and an antiserum against dynorphin(1-13) inhibited clonidine-induced hypotension in spontaneously hypertensive rats, whereas a [Met5]enkephalin antiserum had no effect. Use of antisera specifically recognizing the C-terminus of beta-, alpha-, and gamma-endorphin, respectively, revealed that only the beta-endorphin antiserum inhibited the fall in blood pressure in spontaneously hypertensive rats after cumulative administration of clonidine. None of the antisera used affected clonidine-induced bradycardia. These results indicate that activation of stereospecific opiate receptors in spontaneously hypertensive, but not in Wistar-Kyoto rats, plays a role in the central hypotensive effect of clonidine. beta-Endorphin(1-31) and dynorphin(1-13) might be the endogenous ligands for these receptors.

Absence of brain opioid peptide modulation of luteinizing hormone secretion in the prepubertal gilt.

Three experiments were conducted to evaluate the role of endogenous opioid peptides (EOP) in modulating luteinizing hormone (LH) secretion in the prepubertal gilt. In Experiment I, 8 prepubertal (P) gilts, 160-170 days of age (puberty = 197 +/- 10 days), received either 1 (n = 2), 3 (n = 3), or 6 (n = 3) mg/kg BW of naloxone (NAL), an opiate antagonist, in saline i.v. Blood was collected by jugular vein cannula every 15 min for 2 h before and 2 h after NAL. All doses of NAL failed to alter serum LH concentrations. In Experiment II, 21 P gilts 160-170 days of age and 21 mature (M) gilts were ovariectomized (OVX). At the time of OVX, gilts were classified as prepubertal if their ovaries were devoid of corpora albicantia and corpora lutea. Three weeks after OVX, P and M gilts were injected twice daily for 10 days with either 0.85 mg/kg BW of progesterone (P4) or oil vehicle (V), resulting in the following groups: PP4 (n = 11), PV (n = 10), MP4 (n = 11), and MV (n = 10). All gilts received 1 mg/kg BW of NAL on the last day of treatment. Blood samples were collected via a jugular cannula every 15 min for 4 h before and 2 h after NAL treatment. NAL treatment resulted in an increase (p less than 0.05) in serum LH concentrations only in the MP4 gilts. In Experiment III, 15 OVX gilts 280 days of age were used. Ten of the 15 gilts were OVX prior to puberty at 160 days of age and were classified as chronologically mature (CM) at the time of treatment. The remaining 5 gilts were OVX after puberty, and were classified as sexually mature (SM) at the time of treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Changes in naloxone and haloperidol effects during the development of captivity-induced jumping stereotypy in bank voles

Captivity-induced stereotypies in bank voles ( Clethrionomys glareolus) were inhibited at the age of 1, 2, 4, 6, 8 and 10 months by 0.2 mg/kg haloperidol s.c., while 35 mg/kg naloxone s.c. only inhibited them at the age of 4–6 months. Other activities were not significantly inhibited. The change in the naloxone effect was not due to desensitization as animals treated only at 6 or 8 months reacted similarly. It is suggested that the expression of such stereotypies is influenced by endogenous brain opioid peptides during an early stage of their development, but that thereafter only motoric automatisms remain, essentially under dopaminergic control.

Elevation of brain and adrenal immunoreactive opioid peptide levels in rats during adaptation to physical exercise

The preincubation of isolated washed human platelets with lipopolysaccharide toxin (LPS) from Salmonella typhimurium (10 min, 37 degrees) speeds up the thrombin-induced aggregation. Besides, LPS-pretreatment was shown to alter the turnover of polyphosphoinositides stimulated by thrombin and significantly elevated concentrations of diacylglycerol and thromboxane B2 in comparison with control platelets. However, LPS does not influence on the lipid fluidizing effect of thrombin action as testifies by ESR spectroscopy with doxyl-stearic acids as spin-probes. An LPS pretreatment of platelets that induces no aggregation by itself is proposed might transform cells into a state of hidden activation" associated with the accumulation of such products as diacylglycerol and precursors of thromboxane B2. Addition of thrombin to such "preactivated" platelets could cause a more effective aggregation.

Enkephalinase inhibition: Regulation of ethanol intake in genetically predisposed mice

This is the first report of alteration in alcohol intake in mice with a genetic predisposition to alcohol preference and known to have innate brain enkephalin deficiencies. We have been able to significantly attenuate both volitional and forced ethanol intake respectively by acute and chronic treatment with hydrocinnamic acid and D-phenylalanine, known carboxypeptidase (enkephalinase) inhibitors. Since these agents, through their enkephalinase inhibitory activity, raise brain enkephalin levels, we propose that excessive alcohol intake can be regulated by alteration of endogenous brain opioid peptides.

Effect of central naloxone on hormone and blood pressure responses to hemorrhage in conscious sheep

The role of the brain opioid system in the control of hypothalamic-pituitary-adrenal activity was studied in 10 conscious sheep with an indwelling cannula in a cerebral lateral ventricle. On separate days, sheep received infusions of artificial CSF (control) and the opiate antagonist, naloxone (100 μg/hr) before and during acute moderate hemorrhage (15 ml/kg over 10 min). Infusion of naloxone before hemorrhage raised plasma ACTH and resulted in a significant increase in cortisol responses to hemorrhage control infusion. In contrast, ACTH and cortisol responses to hemorrhage tended to be blunted by central naloxone infusion. The responses of vasopressin, aldosterone and the catecholamines remained unaffected by naloxone. The fall in blood pressure and the rise in heart rate accompanying hemorrhage were likewise unaltered. These results suggest that brain opioid peptides have an inhibitory effect on basal ACTH secretion but do not play a major role in modulating the hemodynamic or pituitary-adrenal responses to acute moderate hemorrhage in conscious sheep. Numerous studies have established that the brain opioid system is activated by a variety of acute stressors, including hemorrhage (1) and other forms of shock and trauma (2, 3). These same stresses are potent stimuli to ACTH secretion. Several lines of evidence indicate that enkephalins and endorphins may participate directly in the regulation of ACTH secretion. The enkephalin agonist, DAMME, inhibits ACTH in sheep (4) and humans (5), opioid peptides suppress corticotropin-releasing factor (CRF) in vitro (6), and the opiate antagonist naloxone enhances ACTH secretion (7), including the ACTH response to exogenous CRF (8). These findings suggest that the brain opioid system may act to restrain both CRF release and ACTH responsiveness during acute stress. Using conscious sheep with indwelling cannulae in a lateral ventricle, we have examined the effect of central naloxone on the ACTH and neuroendocrine response to acute moderate hemorrhage. Since cardio-depressant effects of brain opioid activation also occur during hemorrhage (2, 9) apparently contributing to the shock syndrome, the hemodynamic response to acute, moderate hemorrhage during central administration of naloxone was also studied.