Administration Of Cocaine- And Amphetamine-regulated Transcript Research Articles

A New Insight into the Role of CART Peptide in Serotonergic Function and Anxiety.

Cocaine- and amphetamine-regulated transcript (CART) peptide has been implicated in stress-related behaviors that are regulated by central serotonergic (5-HT) systems in the dorsal raphe nucleus (DRN). Here, we aimed to investigate the interaction between CART and DRN 5-HTergic systems after initially observing CART axonal terminals in the DRN. We found that microinfusion of CART peptide (55-102) into the DRN-induced anxiogenic effects in male C57BL/6J mice, while central administration of CART reduced c-Fos in 5-HTDRN neurons. This inhibitory effect of exogenous CART on 5-HTDRN activity and local 5-HT release was also demonstrated via in vivo fiber photometry coupled with calcium and 5-HT biosensors. CART inputs to the DRN were observed in various subcortical nuclei, but only those in the centrally projecting Edinger-Westphal nucleus (EWcp) were highly responsive to stress. Chemogenetic activation of these DRN-projecting CARTEWcp neurons recapitulated the effects of intra-DRN CART infusion on anxiety-like behavior in males, but not in females, suggesting a sex-specific role for this pathway. Interestingly, CARTEWcp projections to the DRN made direct synaptic contact primarily with non-5-HT neurons, which were also found to express putative CART receptors. Furthermore, chemogenetic stimulation of this CARTEWcp→DRN pathway inhibited 5-HT neurons while increasing activity in local GABAergic neurons. In summary, this study establishes for the first time a neuromodulatory role for CARTEWcp neurons in 5-HTDRN neurotransmission and suggests that CART may drive anxiety-like behavior by promoting feedforward inhibition of 5-HT neurons.

Action of cocaine- and amphetamine-regulated transcript (CART) peptide to attenuate cisplatin-induced emesis in Suncus murinus (house musk shrew)

Cocaine- and amphetamine-regulated transcript (CART) peptide is a brain-gut neuropeptide that has been implicated in a range of physiological functions including appetite, which is disturbed during chemotherapy. The aims of the present study were to identify the distribution and expression of CART mRNA and CART peptide, and to examine the potential of CART (55–102) to attenuate cisplatin-induced emesis in Suncus murinus. CART mRNA and peptide were detected throughout the entire brain, including the forebrain, hypothalamus, and brainstem, and also in the gut wall and stomach. In conscious, freely moving animals, intracerebroventricular administration of CART (55–102) did not modulate food and water intake or alter locomotor activity when administered alone. Cisplatin induced emesis and upregulated the expression of CART mRNA in the brainstem. However, CART (55–102) reduced the number of cisplatin-induced retches. Both CART (55–102) and cisplatin increased the number of c-Fos positive cells in the nucleus tractus solitarius, lateral hypothalamus, paraventricular hypothalamus, and bed nucleus of the stria terminalis (BNST), compared to saline-treated animals, whereas cisplatin also induced c-Fos expression in the area postrema (AP), arcuate nucleus, and central nucleus of the amygdala. Pre-treatment with CART (55–102) significantly attenuated the increased c-Fos positive cells in the BNST and AP. These data indicate that CART mRNA and peptide were localized to regions involved in reward/enforcement, emotion, feeding and emesis. The anti-emetic effect of CART (55–102) against cisplatin-induced emesis may involve both the forebrain limbic system and the brainstem.

Involvement of neuropeptide CART in the central effects of insulin on feeding and body weight

While insulin secreted from pancreas plays a pivotal role in the control of glucose homeostasis, it also interacts with hypothalamic sites and negatively influences the energy balance. The present study was undertaken to reveal the functional interaction between cocaine- and amphetamine-regulated transcript (CART), a well-known anorexic peptide, and insulin within the framework of hypothalamus in the regulation of feeding behavior and body weight. Insulin was administered daily by intracerebroventricular (icv) route, alone or in combination with CART (icv) for a period of seven days. Immediately thereafter, preweighed food was offered to the animals at the commencement of the dark phase. The food intake and body weight were measured daily just prior to next injection. Furthermore, brains of insulin-treated rats were processed for the immunohistochemical analysis of CART-containing elements in the hypothalamus. Treatment with insulin (6 mU, icv) for a period of 7 days caused a significant decrease in food intake and body weight as compared to control. Concomitant administration of CART (0.5 μg, icv) potentiated insulin-induced anorexia and weight loss. Insulin administration resulted in a significant increase in CART immunoreactivity in the hypothalamic arcuate, paraventricular, dorsomedial and ventromedial nuclei. We suggest that increased CART contents in the hypothalamus may be causally linked with anorexia and weight loss induced by insulin.

Central action of CART induces neuronal activation in the paraventricular and dorsomedial hypothalamus of diet-induced obese and lean mice

The cocaine- and amphetamine-regulated transcript (CART) is a peptide commonly studied in the feeding behavior, but it exerts an important role in the autonomic and cardiovascular control as well. It is known that exogenous administration of CART in the central nervous system can elicit increase in blood pressure of both conscious and anesthetized rodents, but little is known whether these central effects might differ between lean and obese animals. We have recently shown that diet-induced obese mice that developed hypertension presented an upregulation of CART levels in the dorsomedial nucleus of hypothalamus, while obese normotensive ones had not. Herein we investigate whether the central action of CART could activate differently the hypothalamic nuclei of diet-induced obese mice compared to the lean counterparts by using Fos protein expression, C57BL/6 mice were randomly assigned to two cohorts, one fed with a high-fat diet for 8 weeks (obese), and the other fed with regular rodent chow (lean). Both groups received an ICV injection of CART at the dose of 400μM, 1 mM or vehicle. Subsequently, the brains were processed for Fos protein immunohistochemical in order to identify hypothalamic neuronal activation. Significantly greater numbers of Fos-positive neurons were observed in the PVN and DMH of obese mice that received CART 1 mM, when compared to the lean control. These results indicate that the central action of CART induces neuronal activation in the hypothalamic nuclei of obese and lean mice, and this could be relevant to the different autonomic and cardiovascular adjustments that an organism exposed to different diet and metabolic condition.

A role of neuropeptide CART in hyperphagia and weight gain induced by olanzapine treatment in rats

Although olanzapine is highly efficacious and most widely used second generation antipsychotic drug, the success of treatment has been hampered by its propensity to induce weight gain. While the underlying neuronal mechanisms are unclear, their elucidation may help to target alternative pathways regulating energy balance. The present study was undertaken to define the role of cocaine- and amphetamine-regulated transcript (CART), a well-known anorexic peptide, in olanzapine-induced hyperphagia and body weight gain in female rats. Olanzapine was administered daily by intraperitoneal route, alone or in combination with CART (intracerebroventricular) for a period of two weeks. Immediately after drug administrations, preweighed food was offered to the animals at the commencement of the dark phase. The food intake and body weight were measured daily just prior to next injection. Furthermore, the brains of olanzapine-treated rats were processed for the immunohistochemical analysis of CART-containing elements in the hypothalamus. Treatment with olanzapine (0.5 mg/kg) for the duration of 14 days produced a significant increase in food intake and body weight as compared to control. However, concomitant administration of CART (0.5 µg) attenuated the olanzapine-induced hyperphagia and weight gain. Olanzapine administration resulted in a significant reduction in CART immunoreactivity in the hypothalamic arcuate, paraventricular, dorsomedial and ventromedial nuclei. We suggest that decreased CART contents in the hypothalamus may be causally linked with the hyperphagia and weight gain induced by olanzapine.

Intraperitoneal Administration of CART 55-102 Inhibits Psychostimulant-Induced Locomotion

CART (cocaine and amphetamine regulated transcript) peptide functions as both a neurotransmitter and a hormone and is found both in the central nervous system (CNS) and in the periphery. CART peptide in the nucleus accumbens (NAc) has been implicated in the regulation of cocaine-dopamine-mediated locomotion and self-administration, and amphetamine-mediated locomotion and behavior. However, there are no studies on the effect of systemic administration of CART peptide on cocaine and amphetamine-mediated locomotion. In this study, we tested if the systemic administration of CART 55-102 by the intraperitoneal (ip) route has a functional effect on psychostimulant-mediated locomotion in rats as it does when given into the brain. We determined that ip CART 55-102 attenuates psychostimulant-mediated locomotion as it does when administered into the NAc and display a biphasic dose response curve.

Cocaine- and Amphetamine-Regulated Transcript (CART) Signaling within the Paraventricular Thalamus Modulates Cocaine-Seeking Behaviour

BackgroundCocaine- and amphetamine-regulated transcript (CART) has been demonstrated to play a role in regulating the rewarding and reinforcing effects of various drugs of abuse. A recent study demonstrated that i.c.v. administration of CART negatively modulates reinstatement of alcohol seeking, however, the site(s) of action remains unclear. We investigated the paraventricular thalamus (PVT) as a potential site of relapse-relevant CART signaling, as this region is known to receive dense innervation from CART-containing hypothalamic cells and to project to a number of regions known to be involved in mediating reinstatement, including the nucleus accumbens (NAC), medial prefrontal cortex (mPFC) and basolateral amygdala (BLA).Methodology/Principal FindingsMale rats were trained to self-administer cocaine before being extinguished to a set criterion. One day following extinction, animals received intra-PVT infusions of saline, tetrodotoxin (TTX; 2.5 ng), CART (0.625 µg or 2.5 µg) or no injection, followed by a cocaine prime (10 mg/kg, i.p.). Animals were then tested under extinction conditions for one hour. Treatment with either TTX or CART resulted in a significant attenuation of drug-seeking behaviour following cocaine-prime, with the 2.5 µg dose of CART having the greatest effect. This effect was specific to the PVT region, as misplaced injections of both TTX and CART resulted in responding that was identical to controls.Conclusions/SignificanceWe show for the first time that CART signaling within the PVT acts to inhibit drug-primed reinstatement of cocaine seeking behaviour, presumably by negatively modulating PVT efferents that are important for drug seeking, including the NAC, mPFC and BLA. In this way, we identify a possible target for future pharmacological interventions designed to suppress drug seeking.

Cocaine- and Amphetamine-Regulated Transcript Peptide Plays a Role in the Manifestation of Depression: Social Isolation and Olfactory Bulbectomy Models Reveal Unifying Principles

We investigated the effect of cocaine- and amphetamine-regulated transcript (CART) peptide on depression-like behavior in socially isolated and olfactory bulbectomized (OBX) rats. Administration of CART (54-102) into the lateral ventricle (50-100 ng) or central nucleus of amygdala (CeA) (10-20 ng) caused significant decrease in immobility time in the forced swim test (FST) without influencing locomotion, suggesting antidepressant-like effect. Social isolation as well as OBX models were undertaken to produce depression-like conditions. Although isolation reared (6 weeks) rats showed significant increase in immobility time in FST, OBX animals exhibited hyperactivity (increase in the ambulation, rearing, grooming, and defecation scores) on day 14 in the open-field test. The isolation- or OBX-induced depression-like phenotypes were reversed following acute or subchronic treatment of CART, respectively, given via intracerebroventricular and intra-CeA routes. Drastic reduction in CART-immunoreactivity was observed in most cells in the paraventricular (PVN), arcuate and Edinger-Westphal nuclei of the socially isolated and OBX animals. Although the fibers in the PVN showed variable response, those in ARC and prefrontal cortex did not change. The CART-immunoreactive fibers in the locus coeruleus also showed highly significant reduction. However, dramatic increase in CART-immunoreactive fibers was noticed in the CeA in both the experimental models. The response by the cells and fibers in the periventricular area and perifornical nucleus in the OBX and socially isolated rats was variable. The study underscores the possibility that endogenous CART system might play a major role in mediating symptoms of depression.

Overexpression of CART in the PVN Increases Food Intake and Weight Gain in Rats

Cocaine- and amphetamine-regulated transcript (CART) codes for a hypothalamic neuropeptide, CART (55-102), which inhibits food intake. Intracerebroventricular injection of CART (55-102) reduces appetite, but also results in motor abnormalities. More recently, studies have demonstrated that administration of CART directly into the paraventricular nucleus (PVN) increases food intake. To investigate the role of CART in the regulation of energy balance in the PVN, we used recombinant adeno-associated virus (rAAV) to overexpress CART in the PVN. Male Wistar rats were injected with either rAAV-encoding CART (rAAV-CART) or rAAV-encoding enhanced green fluorescent protein (rAAV-EGFP) as a control. Food intake and body weight were measured regularly. Animals were fed on normal-chow diet for the first 93 days of the study. After this point, they were fed on high-fat diet. Animals were killed 138 days postinjection and blood and tissues were collected for analysis. Overexpression of CART in the PVN resulted in increased cumulative food intake and body weight gain compared with rAAV-EGFP controls when fed normal chow. These changes became significant at day 65 and 79, respectively and were accentuated on a high-fat diet. A 4% increase in food intake was observed in rAAV-CART animals on a normal-chow diet and a 6% increase when fed a high-fat diet. At the end of the study, rAAV-CART-treated animals had higher circulating leptin concentrations in accord with their higher body weight. These data provide further evidence that hypothalamic CART plays an orexigenic role.

High dose CART peptide induces abnormal EEG activity and behavioral seizures

Cocaine- and amphetamine-regulated transcript (CART) peptides are neurotransmitters found throughout the nervous system and in the periphery. CART has an important role in the regulation of food intake, anxiety, endocrine function, and in mesolimbic-mediated reward and reinforcement. This short report casts light upon previous descriptions of presumed behavioral seizure and tremor activity following administration of CART into the central nervous system. By employing electroencephalographic (EEG) recordings, we document the state of cerebrocortical activity. We find that intracerebroventricular (icv) administration of 5 microg of CART 55-102 readily produces an abnormal EEG characterized initially by high amplitude hypersynchronous alpha in the 8-10 Hz range during behavioral wakefulness as manifest in both cortical and hippocampal theta EEG channels. This reliably progressed in three of three animals tested to unequivocal epileptiform activity accompanied by tremors and assumption of a rigid, tonic body posture. The neural substrates underlying this finding are unclear. This novel description of the epileptogenic quality of CART should lend caution to interpretations of the behaviors attributed to CART in other experimental paradigms.

Central administration of insulin suppresses food intake in chicks

Although the orexigenic action of peptide hormones such as ghrelin and growth hormone releasing peptide is different between chickens and mammals, the anorexigenic action of peptide hormones is similar in both species. For example, central administration of peptide hormones such as leptin, cholecystokinin or glucagon has been shown to suppress food intake behavior in chickens and mammals. Central administration of insulin suppresses food intake in mammals. However, the anorexigenic action of insulin in chickens has not yet been identified. In the present study, we investigated the effects of central administration of insulin on food intake in chicks. Intracerebroventricular administration of insulin in chicks significantly suppressed food intake. Central administration of insulin significantly upregulated mRNA levels of proopiomelanocortin (POMC), cocaine- and amphetamine-regulated transcript (CART) and corticotropin-releasing factor (CRF), but did not influence mRNA levels of neuropeptide Y and agouti-related protein in the hypothalamus. These results suggest that alpha-melanocyte stimulating hormone (alpha-MSH, an anorexigenic peptide from the post-translational cleavage of POMC), CART and CRF are involved in the anorexigenic action of insulin in chicks. Furthermore, central administration of alpha-MSH or CART significantly suppressed food intake. In addition, alpha-MSH significantly upregulated CRF mRNA expression, suggesting that the anorexigenic action of alpha-MSH is mediated by CRF. Our findings demonstrate that insulin functions in chicks as an appetite-suppressive peptide in the central nervous system and suggest that this anorexigenic action is mediated by CART, alpha-MSH and CRF.

Dissecting the role of cocaine- and amphetamine-regulated transcript (CART) in the control of appetite

Cocaine- and amphetamine-regulated transcript (CART) codes for a neuropeptide system with a number of biological roles. The high conservation of CART across species suggests that it has an important role in mammalian physiology. CART is widely expressed in the central nervous system and the periphery, but is particularly concentrated in the hypothalamus. CART peptides, particularly CART (55-102), appear to have an important function in the regulation of energy homeostasis. This review aims to dissect the role of CART in appetite and energy expenditure. CART interacts with a number of central appetite circuits. Hypothalamic CART expression is regulated by a number of peripheral factors, including the adipose hormone leptin. Intracerebroventricular administration of CART (55-102) reduces appetite and stimulates energy expenditure. Hypothalamic CART may also play an orexigenic role under specific circumstances, however, as injection of CART (55-102) into specific hypothalamic nuclei increases food intake.

Intrathecal CART (55-102) enhances the spinal analgesic actions of morphine in mice

Cocaine- and amphetamine-regulated transcript (CART) peptides are found in brain and spinal cord areas involved in pain transmission. In the present study, we investigated the role of rat CART (55-102) in the modulation of an acute pain model after intrathecal administration. The results show that CART (55-102) was without effect on the tail-flick test after i.t. injection in mice. Interestingly, i.t. administration of CART (55-102) significantly enhanced the antinociceptive effect of morphine in the tail-flick test. These results suggest that CART (55-102) potentiates the effects of opioids to inhibit the nociceptive information transmission.

Central administration of cocaine- and amphetamine-regulated transcript increases phosphorylation of cAMP response element binding protein in corticotropin-releasing hormone-producing neurons but not in prothyrotropin-releasing hormone-producing neurons in the hypothalamic paraventricular nucleus

Cocaine- and amphetamine-regulated transcript (CART) has an important action on hypophysiotropic thyrotropin-releasing hormone (TRH) and corticotropin-releasing hormone (CRH) neurons to regulate the hypothalamic–pituitary–thyroid and adrenal axis, respectively. To elucidate the mechanisms by which CART mediates its effect on TRH and CRH neurons, we determined whether the exogenous administration of CART into the cerebrospinal fluid (CSF) phosphorylates the transcription factor, cyclic adenosine 5′-monophosphate response element binding protein (CREB), in the nucleus of TRH and CRH neurons. CART dramatically increased the percentage of phosphoCREB (PCREB) immunolabeled cell nuclei in the hypothalamic paraventricular nucleus (PVN) in fasted as well as fed rats at 10-min postinjection, particularly in the medial parvocellular subdivision of the PVN. Double immunolabelling with CRH antiserum revealed that CART increased the number of CRH neurons containing PCREB from 10.5±1.2 % to 87±1.2% ( P<0.001) in fasting animals and from 3.7±0.8% to 74±5.3% ( P<0.001) in fed animals. In contrast, no significant change was observed in the percentage of proTRH neurons colocalizing with PCREB either in the fasted (11.7±1.85%) or fed animals (4.2±2.2%) as compared to their respective vehicle controls (2.5±1.4% and 4.6±1%). Ultrastructural analysis revealed that CART establishes axosomatic and axodendritic contacts with CRH neurons in the PVN. These data demonstrate a selective effect of CART to phosphorylate CREB in CRH, but not TRH neurons in the PVN. Since CART is capable of increasing the gene expression of both CRH and TRH in hypophysiotropic neurons, and CART-containing axon terminals establish synaptic relationships with hypophysiotropic CRH and TRH neurons, we propose that CART may signal to the nucleus by more than one pathway.

Cocaine- and amphetamine-regulated transcript peptide produces anxiety-like behavior in rodents

Cocaine- and amphetamine-regulated transcript (CART) peptide (CART-(55–102)) is involved in the suppression of food intake. We now report that CART-(55–102) is involved in anxiety in rodents. Intracerebroventricularly administered CART-(55–102) as well as intraperitoneal administration of N-methyl-β-carboline-3-carboxamide (FG-7142), a selective GABA A/benzodiazepine receptor inverse agonist, reduced time spent in the open arms in the elevated plus-maze task in mice. CART-(55–102)-induced anxiogenic-like behavior in this task was attenuated by widely prescribed anxiolytics such as diazepam and buspirone. Likewise, CART-(55–102) and FG-7142 significantly reduced social interaction in mice. Both diazepam and buspirone significantly reversed CART-(55–102)-induced anxiogenic-like behavior in social interaction tests. By contrast, another biologically active CART peptide, CART-(62–102), was without effect in the elevated plus-maze task in mice. Moreover, intracerebroventricular administration of CART-(55–102) markedly increased the firing rate of locus coeruleus neurons in single unit recording in anesthetized rats. As CART-(55–102) produced anxiety-like effects in rodents, this peptide may possibly be involved in anxiety and stress-related behavior.

Anorexigenic cocaine- and amphetamine-regulated transcript peptide intensifies fear reactions in rats

An increasing number of appetite-regulating peptides are being discovered. The list of regulators inhibiting food intake is considerably longer than that of appetite stimulators. In many cases, the peptides inhibiting food intake facilitate fear reactions, whereas the majority of the agents reducing anxiety responses stimulate appetite. Cocaine- and amphetamine-regulated transcript (CART) cDNA was isolated from hypothalamic libraries and CART was reported to inhibit food intake and to mediate the anorectic effects of leptin. Here, we show that the active core fragment of CART (CART 89–103, 0.04–5.0 nmol) injected into lateral cerebral ventricle not only inhibits food intake, but also causes a dose-dependent increase in anxiety-like reactions in elevated plus-maze test. Intracerebroventricular administration of CART 82–103 (0.04–5.0 nmol) did not inhibit water intake and did not affect spontaneous locomotor activity in the open field test ruling out unspecific effects of the peptide. Our results suggest that CART could be an endogenous factor in the brain mediating the effects of stress on appetite.

The hypothalamic satiety peptide CART is expressed in anorectic and non-anorectic pancreatic islet tumors and in the normal islet of Langerhans

The hypothalamic satiety peptide CART (cocaine and amphetamine regulated transcript) is expressed at high levels in anorectic rat glucagonomas but not in hypoglycemic insulinomas. However, a non-anorectic metastasis derived from the glucagonoma retained high CART expression levels and produced circulating CART levels comparable to that of the anorectic tumors. Moreover, distinct glucagonoma lines derived by stable HES-1 transfection of the insulinoma caused severe anorexia but retained low circulating levels of CART comparable to that of insulinoma bearing or control rats. Islet tumor associated anorexia and circulating CART levels are thus not correlated, and in line with this peripheral administration of CART (5–50 mg/kg) produced no effect on feeding behavior. In the rat two alternatively spliced forms of CART mRNA exist and quantitative PCR revealed expression of both forms in the hypothalamus, in the different islet tumors, and in the islets of Langerhans. Immunocytochemistry as well as in situ hybridization localized CART expression to the somatostatin producing islet D cell. A potential endocrine/paracrine role of islet CART remains to be clarified.