Endocrine Abstracts (2003) 5 OC32

Localisation and regulation of cocaine- and amphetamine-regulated transcript (CART) in the anterior pituitary gland

KG Murphy, SA Stanley, G Bewick, WM Kong, JV Gardiner, M Ghatei, CJ Small & SR Bloom

Endocrine Unit, Imperial College School of Science, Technology and Medicine, London, UK.

Cocaine and amphetamine regulated transcript (CART) was originally isolated from rat brain but CART is also synthesized and stored in the anterior pituitary. The localisation and factors regulating pituitary CART synthesis are largely unknown.
Using a specific assay for the pituitary form of CART, CART(55-102), CART-immunoreactivity was detectable in the medium of the corticotroph cell line, AtT-20 (1.73 ± 0.54 fmols per millilitre per 24h) but not in control medium. Dual in situ hybridisation for preproCART (ppCART) mRNA expression and immunocytochemistry for ACTH-ir showed co-localisation of ppCART mRNA to a subpopulation of ACTH immunoreactive cells.
We went on to determine if pituitary CART expression and release are also regulated by corticotrophin releasing factor (CRF). Short-term incubation of pituitary segments with CRF (100 nanomolar) increased CART-ir release by fifteen fold. Adrenalectomy and excess glucocorticoid treatment result in long-term alternations in CRF and ACTH. Pituitary CART mRNA expression, pituitary CART-ir content and plasma CART-ir were measured in sham-operated, adrenalectomised and corticosterone-treated rats. Pituitary CART mRNA, CART-ir content and circulating CART-ir were all significantly increased in adrenalectomised rats and decreased in corticosterone-treated rats.
Finally, to determine if CART might play a paracrine role in the pituitary, the effect of CART(55-102) on ACTH release from pituitary segments was examined. CART(55-102) significantly increased basal and CRF-stimulated ACTH release (ACTH release: basal 100 ± 6 % basal, CART (55-102) 100nM 127 ± 9 % basal, p less than 0.05, CRF 100nM 289 ± 17 % basal, CRF 100nM + CART (55-102) 10nM 391 ± 31 % basal, p less than 0.05).
These findings suggest pituitary CART is partly co-localised with pituitary ACTH. Furthermore, pituitary CART expression, content and release are regulated by hypothalamic CRF and CART increases pituitary ACTH release. These findings suggest pituitary CART may also play a role in regulation of the stress response.

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