Research ReportCocaine- and amphetamine-regulated transcript (CART) peptide- immunopositive neuronal elements in the lateral septum: Rostrocaudal distribution in the male rat
Research highlights
►CART-IR is present along the rostrocaudal axis of the rat LS in varicose nerve fibers. ►Two morphologically distinct types of fibers exhibited CART-IR. ►CART-IR around GAD- and NPY-containing cell bodies was revealed. ►Smaller boutons gave asymmetrical, while larger boutons formed symmetrical synapses. ►Densitometry of serial sections revealed a paraboloid distribution of CART.
Introduction
CART (cocaine- and amphetamine-regulated transcript) peptides were shown to be involved in a range of physiological processes such as feeding, locomotion, drug reward, reinforcement, and stress (Hunter and Kuhar, 2003). CART was first identified as an upregulated transcript in the rat nucleus accumbens following the acute administration of psychostimulants, cocaine and amphetamine (Douglass et al., 1995). This mRNA encodes a precursor peptide of either 116 or 129 amino acids. After the cleavage of some amino acids either the long form (102 amino acids) or the short form (89 amino acids) represents the active transmitter molecules. It was suggested that CART might be involved in feeding, since CART immunoreactivity was localized in brain regions associated with the regulation of food intake viz. the lateral hypothalamus (LH), ventromedial nucleus (VMN), dorsomedial nucleus (DMN), arcuate nucleus (Arc), paraventricular nucleus (PVN) of the hypothalamus, and nucleus accumbens (Koylu et al., 1997, Koylu et al., 1998). CART is expressed in several other brain regions, including the pituitary gland, nuclei of amygdala, olfactory bulb, Edinger–Westphal nucleus, and nuclei of the solitary tract (Koylu et al., 1998, Hurd and Fagergren, 2000, Kozicz, 2003). CART was also found at several peripheral locations that are involved in feeding, including the adrenal medulla, stomach, duodenum, the myenteric plexus of the gastrointestinal tract, and vagus nerve (Koylu et al., 1997, Koylu et al., 1998, Couceyro et al., 1998, Broberger et al., 1999, Kuhar and Yoho, 1999, Ekblad et al., 2003).
In the central nervous system, CART peptides are colocalized with other neurotransmitters and peptides that are also involved in the regulation of feeding, such as melanin-concentrating hormone in the DMN and LH (Broberger, 1999) and pro-opiomelanocortin (α-melanocyte-stimulating hormone) in the Arc (Elias et al., 1998). There is a functional interaction between CART peptide and the strongly orexigenic neuropeptide Y (NPY), since NPY-positive varicosities were observed around CART peptide-positive cell bodies in the PVN (Lambert et al., 1998). Kristensen et al. (1998) demonstrated that CART is a potent anorexigenic peptide. More specifically, a) intracerebroventricular administration of CART peptide fragments inhibits both normal and starvation-induced feeding in rats and completely blocks the feeding response induced by neuropeptide Y (Kristensen et al., 1998) and b) leptin receptors are present in CART-IR neurons in the Arc and leptin has been shown to be one of the strongest regulators of CART mRNA levels in the hypothalamus (Elias et al., 1998, Kristensen et al., 1998).
Previous studies indicated that the lateral septal area is rich in neuropeptides involved in the regulation of feeding such as galanin (gal), NPY, and opioids (Kovács et al., 2005); moreover, they respond with density changes to food deprivation (Kovács et al., 2007). The aims of the present study were to determine the presence of CART peptide in the LS, to describe the light and electron microscopic morphology of neuronal elements expressing the CART peptide, to reveal their fine structural characteristics and synaptic targets, and to determine the rostrocaudal distribution of CART in the lateral septum of male rats.
Section snippets
Results
CART-immunoreactive axonal fibers were detected along the rostrocaudal axis of the LS (Fig. 1). The immunoprecipitate was present in varicose nerve fibers; however, CART-IR was observed neither in local somata nor in dendrites. The dense immunoreactive plexus was not restricted to the lateral part of the septal complex; it also extended to the medial septum–diagonal band of Broca (MS/DB).
Discussion
This work provides immunocytochemical evidence on the presence of the CART peptide in the LS of male rats, and the major findings of this study are as follows: a) two types of CART-IR axon terminals were demonstrated with distinct distributional pattern, b) the distribution of CART-IR nerve plexuses showed a parabolic curve, and c) most of the synapses formed by CART-IR neuronal structures were asymmetric.
Animals and experimental groups
The experiments were carried out on 14 young adult male Wistar rats of the same age weighing 230–250 g. All animal procedures were conducted in accordance with the guidelines set forth by the European Communities Council Directive of 24 November 1986 (86/609/EEC) and the Animal Health and Welfare Institute of the Szent István University, and all efforts were made to minimize the number of animals and to keep animal stress, suffering, and discomfort to a minimum level. The Local Animal Welfare
Acknowledgments
The authors acknowledge the support of Hungarian Research FundOTKA T 43170 and are grateful for Dr. J.T. Clausen for the donation of the monoclonal CART antibody.
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