Quantification and synthesis of cocaine- and amphetamine-regulated transcript peptide (79-102)-like immunoreactivity and mRNA in rat tissues

Abstract

The distribution of cocaine- and amphetamine-regulated transcript peptide (79-102)-like immunoreactivity (CART-LI) was quantified in brain and peripheral tissues of male and female Wistar rats, and male obese (fa/fa) and heterozygous (Fa/+) Zucker rats using a specific RIA. CART-LI tissue levels have not been quantified previously. The assay, using cocaine- and amphetamine-regulated transcript (CART) (79-102) as a standard and radioactive tracer and an antibody to CART (79-102) fragment, detected CART-LI in all brain regions examined, the anterior and posterior pituitary, the spinal cord and throughout the gastrointestinal tract of both male and female Wistar rats. The highest concentrations were found in the hypothalamus, duodenum, anterior pituitary and posterior pituitary (50.6 ± 4.4, 26.1 ± 4.2, 50.0 ± 1.3 and 373.0 ± 55.2 pmol/g wet tissue respectively, means ± S.E.M., n=6-10 male animals). There was no significant variation between the sexes. The concentrations of CART-LI in hypothalami and anterior and posterior pituitaries from fa/fa rats were significantly (P<0002) lower than those of Fa/+ controls (35.9 ± 2.1 vs 53.9 ± 4.9, <0.6 vs 1.8 ± 0.4 and 114 ± 9.1 vs 255.5 ± 20.9 pmol/g wet tissue respectively, means ± S.E.M., n=7). Gel permeation chromatography of regions of rat brain and gastrointestinal tract showed possible differential processing between regions. CART-LI was released from hypothalamic tissue slices in a calcium-dependent fashion by potassium-induced depolarisation. Northern blot analysis detected CART mRNA in the hypothalamus, anterior pituitary, brain stem, cerebellum and spinal cord. The pattern of distribution of CART mRNA and CART-LI in various neural and other tissues is in accord with a role for CART as a neurotransmitter.