Proceedings of The Physiological Society

University of Bristol (2005) J Physiol 567P, PC55

Poster Communications

Visualisation of hindbrain target neurones for membrane actions of corticosterone

Rose, James D.; Dolence, Kurt; Mitchell, Jeanette A.; Coddington, Emma J.; Hubbard, Catherine S.;

1. Zoology/Physiology , University of Wyoming, Laramie, WY, USA. 2. School of Pharmacy, University of Wyoming, Laramie, WY, USA.

A specific, high-affinity neuronal membrane receptor for corticosterone (CORT), with binding properties distinct from intracellular glucocorticoid receptors, mediates rapid suppressive effects of stress and CORT secretion on courtship clasping behaviour in the roughskin newt (Taricha granulosa). Our previous studies showed that CORT acts through this membrane receptor to rapidly disrupt clasping by depressing medullary reticular neurone responsiveness to clasp-triggering sensory stimuli (e.g. pressure on the cloaca) and by interfering with the functioning of reticulospinal neurones. While medullary reticular neurones are affected by CORT action, the identity of specific neurones possessing functional CORT membrane receptors is unknown. To address this question, we prepared a conjugate of CORT and the fluorescent dye, Cascade Blue (CB). This conjugate was injected systemically in unanaesthetized newts or applied directly to the medullary surface of newts anaesthetized by immersion in 0.1% MS-222. Reticulospinal neurones were retrogradely labelled with tetramethylrhodamine dextran-amine administered to a hemisected spinal cord at the first cervical vertebral level under MS-222 anaesthesia. Epifluorescent microscopic examination of medullary brain sections revealed CORT-CB internalisation as granular fluorescence in the soma and proximal dendrites of a large proportion of neurones. These neurones were widely distributed in the medulla and constituted a diversity of neuronal phenotypes. CORT-CB internalisation was most prominent 30 min after application, and was blocked by pretreatment with unlabelled CORT as well as the kappa-opiate receptor agonist, U-50488, but not by dexamethasone (DEX), results that match the known binding pattern of the CORT membrane receptor. Systemic administration or medullary CORT-CB application rapidly altered neuronal activity and sensory responsiveness similar to effects of unconjugated CORT. Conjugate administration also produced non-granular, nuclear labelling in some neurones, apparently due to intracellular glucocorticoid receptor binding because it was specifically blocked by pretreatment with DEX but not U-50488. This new approach for functional and structural identification of the neurones with CORT membrane receptors should greatly facilitate our understanding of the neural basis for the behavioural effects of the hormone.

Where applicable, experiments conform with Society ethical requirements