A COCAINE-AND-AMPHETAMINE-REGULATED-TRANSCRIPT PEPTIDE PROJECTION FROM THE LATERAL HYPOTHALAMUS TO THE VENTRAL TEGMENTAL AREA K. B. PHILPOT, a1 * S. DALLVECHIA-ADAMS, a1 Y. SMITH a,b2 AND M. J. KUHAR a2 a Yerkes National Primate Research Center, Division of Neuroscience, 954 Gatewood Road Northeast, Atlanta, GA 30329, USA b Department of Neurology, Emory University, 954 Gatewood Road Northeast, Atlanta, GA 30322, USA Abstract—Cocaine-and-amphetamine-regulated-transcript pep- tides play a role in the modulation of feeding and psychomo- tor stimulant-like behaviors. The ventral tegmental area and the lateral hypothalamus are likely structures where cocaine- and-amphetamine-regulated-transcript peptides mediate both of these functions. Although lateral hypothalamus inputs to the ventral tegmental area have long been known, the chemical nature of this pathway remains poorly understood. To address this issue, we tested the possibility that cocaine-and-amphet- amine-regulated-transcript peptide-containing neurons in the lateral hypothalamus project to the ventral tegmental area using the retrograde transport of cholera toxin subunit B combined with cocaine-and-amphetamine-regulated-transcript peptide immunostaining. The largest density of retrogradely-labeled neurons in the hypothalamus after cholera toxin subunit B injection in the ventral tegmental area was found, ipsi- and contralaterally, in the lateral hypothalamus/perifornical area, although substan- tial numbers of retrogradely-labeled cells were also found in the medial preoptic area, lateral preoptic area, paraven- tricular nucleus, dorsomedial hypothalamus and ventrome- dial hypothalamus. More than 80% of the retrogradely-labeled cocaine-and-amphetamine-regulated-transcript peptide-im- munoreactive neurons in the hypothalamus were found in the lateral hypothalamus/perifornical area both ipsilateral and contralateral to the injection sites. Although retrogradely- labeled neurons were seen in the amygdala, locus coeruleus, and raphe nucleus, none of them displayed cocaine-and- amphetamine-regulated-transcript peptide immunoreactivity. Therefore, the hypothalamic projection to the ventral teg- mental area provides a substrate whereby cocaine-and-am- phetamine-regulated-transcript peptides could mediate the rewarding aspects of feeding and psychomotor stimulant-like behaviors. These findings, combined with the fact that the lateral hypothalamus receives strong inputs from the shell of the nucleus accumbens and ventral pallidum, suggest that these structures are part of integrative functional loops that control reward and appetitive behaviors. © 2005 IBRO. Pub- lished by Elsevier Ltd. All rights reserved. Key words: accumbens, cholera toxin, retrograde tracing, psychomotor stimulants, feeding behavior. The midbrain dopaminergic neurons of the ventral tegmen- tal area (VTA) play an important role in mediating the reinforcing properties of drugs of abuse. Many neuropep- tides modulate midbrain dopaminergic activity in the VTA (Kelley and Cador, 1988). Accordingly, studies elucidating the mechanism of action of the neuropeptide, cocaine-and- amphetamine-regulated-transcript (CART), have demon- strated that CART peptides are modulators of mesolimbic dopamine and psychostimulants (Kimmel et al., 2000; Ja- worski et al., 2003a,b; Kim et al., 2003; Shieh, 2003; Yang et al., 2004). Discrete injection of CART peptides into the VTA produces the psychomotor stimulant-like effects of increased locomotor activity and conditioned place prefer- ence in the rat (Kimmel et al., 2000), although such an injection also reduces locomotor effects of cocaine (Jawor- ski et al., 2004). To better understand the mechanisms of action that may underlie CART peptide’s effects in the VTA, we recently examined the synaptology and neuro- chemistry of cocaine-and-amphetamine-regulated-tran- script peptide-immunoreactive (CARTir) terminals in the rat VTA (Dallvechia-Adams et al., 2002). Results of this study showed that CARTir terminals form both symmetric and asymmetric synapses on dopaminergic and non-do- paminergic VTA neurons, indicating that the CARTir affer- ents likely arise from at least two different sources. Based on previous studies, five potential nuclei may provide CARTir afferents to the VTA. These nuclei that project to the VTA (Phillipson, 1979; Zahm et al., 2001) and contain CARTir cell bodies (Koylu et al., 1997; Koylu et al., 1998) include the nucleus accumbens (Acb), amygdala, hypo- thalamus, locus coeruleus (LC) and raphe nucleus (RN). The fact that many CART-containing varicosities in the VTA and CARTir cell bodies in the lateral hypothalamic perifornical area (LH/Pef) co-express melanin-concentrat- ing hormone (MCH) (Broberger, 1999; Vrang et al., 1999; Elias et al., 2001; Dallvechia-Adams et al., 2002) suggests that at least part of the CART innervation of the VTA arises from the LH/Pef. To test this possibility, we placed discrete injections of the retrograde tracer, cholera toxin subunit B 1 These authors contributed equally to the work. 2 These authors contributed equally to the work. *Corresponding author. Tel: +1-404-727-1737; fax: +1-404-727-3278. E-mail address: sevenseat_2000@yahoo.com (K. B. Philpot). Abbreviations: ABC, avidin– biotin peroxidase complex; Acb, nucleus accumbens; AcbSh, nucleus accumbens shell; BSA, bovine serum albumin; CART, cocaine-and-amphetamine-regulated-transcript; CARTir, CART immunoreactive; CTB, cholera toxin subunit B; CTBir, CTB im- munoreactive; DMH, dorsomedial hypothalamus; LC, locus coeruleus; LH/Pef, lateral hypothalamic perifornical area; LPO, lateral preoptic area; MCH, melanin-concentrating hormone; MPO, medial preoptic area; NHS, normal horse serum; NMS, normal mouse serum; NT, neuro- tensin; Orx, orexin; PB, phosphate buffer; PBS, phosphate-buffered saline; PVN, paraventricular nucleus; RN, raphe nucleus; RRF, ret- rorubral field; SN, substantia nigra; VMH, ventromedial hypothalamus; VP, ventral pallidum; VTA, ventral tegmental area. Neuroscience 135 (2005) 915–925 0306-4522/05$30.00+0.00 © 2005 IBRO. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.neuroscience.2005.06.064 915