CART neurons in the lateral hypothalamus communicate with the nucleus accumbens shell via glutamatergic neurons in paraventricular thalamic nucleus to modulate reward behavior.

Paraventricular thalamic nucleus (PVT) serves as a transit node processing food and drug-associated reward information, but its afferents and efferents have not been fully defined. We test the hypothesis that the CART neurons in the lateral hypothalamus (LH) project to the PVT neurons, which in turn communicate via the glutamatergic fibers with the nucleus accumbens shell (AcbSh), the canonical site for reward. Rats conditioned to self-stimulate via an electrode in the right LH-medial forebrain bundle were used. Intra-PVT administration of CART (55-102) dose-dependently (10-50 ng/rat) lowered intracranial self-stimulation (ICSS) threshold and increased lever press activity, suggesting reward-promoting action of the peptide. However, treatment with CART antibody (intra-PVT) or MK-801 (NMDA antagonist, intra-AcbSh) produced opposite effects. A combination of sub-effective dose of MK-801 (0.01 µg/rat, intra-AcbSh) and effective dose of CART (25 ng/rat, intra-PVT) attenuated CART's rewarding action. Further, we screened the LH-PVT-AcbSh circuit for neuroadaptive changes induced by conditioning experience. A more than twofold increase was noticed in the CART mRNA expression in the LH on the side ipsilateral to the implanted electrode for ICSS. In addition, the PVT of conditioned rats showed a distinct increase in the (a) c-Fos expressing cells and CART fiber terminals, and (b) CART and vesicular glutamate transporter 2 immunostained elements. Concomitantly, the AcbSh showed a striking increase in expression of NMDA receptor subunit NR1. We suggest that CART in LH-PVT and glutamate in PVT-AcbSh circuit might support food-seeking behavior under natural conditions and also store reward memory.

CART neuropeptide modulates the extended amygdalar CeA-vBNST circuit to gate expression of innate fear.

Innate fear is critical for the survival of animals and is under tight homeostatic control. Deregulation of innate fear processing is thought to underlie pathological phenotypes including, phobias and panic disorders. Although central processing of conditioned fear has been extensively studied, the circuitry and regulatory mechanisms subserving innate fear remain relatively poorly defined. In this study, we identify cocaine- and amphetamine-regulated transcript (CART) neuropeptide signaling in the central amygdala (CeA) - ventral bed nucleus of stria terminalis (vBNST) axis as a key modulator of innate fear expression. 2,4,5-trimethyl-3-thiazoline (TMT), a component of fox faeces, induces a freezing response whose intensity is regulated by the extent of CART-signaling in the CeA neurons. Abrogation of CART activity in the CeA attenuates the freezing response and reduces activation of vBNST neurons. Conversely, ectopically elevated CART signaling in the CeA potentiates the fear response concomitant with enhanced vBNST activation. We show that local levels of CART signaling modulate the activation of CeA neurons by NMDA receptor-mediated glutamatergic inputs, in turn, regulating activity in the vBNST. This study identifies the extended amygdalar CeA-vBNST circuit as a CART modulated axis encoding innate fear. CART signaling regulates the glutamatergic excitatory drive in the CeA-vBNST circuit, in turn, gating the expression of the freezing response to TMT.

Identification of the CART neuropeptide circuitry processing TMT-induced predator stress.

Abundance of cocaine- and amphetamine-regulated transcript (CART) neuropeptide in the limbic areas like the olfactory system, central nucleus of amygdala (CeA), ventral bed nucleus of stria terminalis (vBNST) and the hypothalamus suggests involvement of the peptide in emotive processing. We examined the role of CART in mediating fear, a strong emotion with profound survival value. Rats, exposed to 2,4,5-trimethyl-3-thiazoline (TMT), a predator related cue extracted from fox feces, showed significant increase in freezing, escape and risk assessment behavior, whereas grooming was reduced. Neuronal activity was up-regulated in the CeA and vBNST in terms of increased immunoreactivity in CART elements and c-Fos expression. Increased expression of both the markers was also seen in some discrete magnocellular as well as parvicellular subdivisions of the paraventricular nucleus (PVN). However, CART containing mitral cells in the main or accessory olfactory bulb did not respond. CART antibody was stereotaxically injected bilaterally into the CeA to locally immunoneutralize endogenous CART. On exposure to TMT, these rats showed reduced freezing, risk assessment and escape behavior while grooming was restored to normal value. We suggest that the CART signaling in the CeA and vBNST, but not in the olfactory system, might be an important component of the innate fear processing, and expression of stereotypic behavior, while CART in the PVN subdivisions might mediate the neuroendocrine response to predator stress.