Association of cocaine- and amphetamine-regulated transcript-immunoreactive elements with thyrotropin-releasing hormone-synthesizing neurons in the hypothalamic paraventricular nucleus and its role in the regulation of the hypothalamic-pituitary-thyroid axis during fasting

Because cocaine- and amphetamine-regulated transcript (CART) coexists with alpha -melanocyte stimulating hormone (alpha -MSH) in the arcuate nucleus n eurons and we have recently demonstrated that alpha -MSH innervates TRH-syn thesizing neurons in the hypothalamic paraventricular nucleus (PVN), we rai sed the possibility that CART may also be contained in fibers that innervat e hypophysiotropic thyrotropin-releasing hormone (TRH) neurons and modulate TRH gene expression. Triple-labeling fluorescent in situ hybridization and immunofluorescence were performed to reveal the morphological relationship s between pro-TRH mRNA-containing neurons and CART- and alpha -MSH-immunore active (IR) axons. CART-IR axons densely innervated the majority of pro-TRH mRNA-containing neurons in all parvocellular subdivisions of the PVN and e stablished asymmetric synaptic specializations with pro-TRH neurons. Howeve r, whereas all alpha -MSH-IR axons in the PVN contained CART-IR, only a por tion of CART-IR axons in contact with pro-TRH neurons were immunoreactive f or alpha -MSH. In the medial and periventricular parvocellular subdivisions of the PVN, CART was co-contained in similar to 80% of pro-TRH neuronal pe rikarya, whereas colocalization with pro-TRH was found in <10% of the anter ior parvocellular subdivision neurons. In addition >80% of TRH/CART neurons in the periventricular and medial parvocellular subdivisions accumulated F luoro-Gold after systemic administration, suggesting that CART may serve as a marker for hypophysiotropic TRH neurons. CART prevented fasting-induced suppression of pro-TRH in the PVN when administered intracerebroventricular ly and increased the content of TRH in hypothalamic cell cultures. These st udies establish an anatomical association between CART and pro-TRH-producin g neurons in the PVN and demonstrate that CART has a stimulatory effect on hypophysiotropic TRH neurons by increasing pro-TRH gene expression and the biosynthesis of TRH.