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Origin of Cocaine- and Amphetamine-Regulated Transcript-Containing Axons Innervating Hypophysiotropic Corticotropin-Releasing Hormone-Synthesizing Neurons in the Rat

Department of Endocrine Neurobiology (G.W., Z.L., C.F.), Institute of Experimental Medicine, Hungarian Academy of Sciences, Budapest 1083, Hungary; and Tupper Research Institute and Department of Medicine (R.M.L., C.F.), Division of Endocrinology, Diabetes, Metabolism, and Molecular Medicine, Tufts-New England Medical Center, and Department of Neuroscience (R.M.L.), Tufts University School of Medicine, Boston, Massachusetts 02111

Address all correspondence and requests for reprints to: Csaba Fekete M.D., Ph.D., Department of Endocrine Neurobiology, Institute of Experimental Medicine, Hungarian Academy of Sciences, Budapest 1083, Hungary. E-mail: feketecs{at}koki.hu.

Cocaine- and amphetamine-regulated transcript (CART) has stimulatory effects on the hypothalamic-pituitary-adrenal axis through direct effects on hypophysiotropic CRH neurons. Recently CART-containing axons have been demonstrated to densely innervate the hypophysiotropic CRH neurons. Based on the sources of the CART-immunoreactive (IR) innervation of the paraventricular nucleus, the putative origins of these CART-containing fibers include neurons of the hypothalamic arcuate nucleus that coexpress MSH and medullary adrenaline-producing neurons. To determine whether these cell groups contribute to the CART innervation of the hypophysiotropic CRH neurons, we performed a quadruple-labeling immunofluorescent study using antisera against CRH, CART, MSH, and phenylethanolamine-N-methyl-transferase (PNMT), the latter as a marker for adrenaline. Consistent with previous observations, PNMT- and CART-IR axons densely innervated all CRH neurons, whereas the MSH-IR innervation was sparse. Although approximately 60% of CART-IR varicosities in juxtaposition to CRH neurons cocontained PNMT, only approximately 18% of them were immunopositive for MSH. All MSH-IR boutons and approximately 90% of PNMT-containing varicosities on the surface of CRH neurons were also labeled for CART. The remaining 22% of CART axon varicosities in contact with CRH neurons contained neither MSH nor PNMT. These results indicate that medullary adrenergic/CART neurons are the major source for the CART innervation of CRH neurons in the paraventricular nucleus; however, to a lesser extent the arcuate nucleus also contributes to the CART-IR innervation of these neurons. The observation that nearly 20% of the CART-IR afferents contain neither MSH nor PNMT, however, suggests that additional sources also contribute to the CART-IR input of hypophysiotropic CRH neurons.

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