In mammals, adrenal zona fasciculata (AZF) cells secrete cortisol which acts on cells of the liver, muscle, and adipose tissue to enhance glucose synthesis and to promote the breakdown of fat and muscle proteins. Precise control of blood glucose concentration by cortisol and other hormones is critical because this sugar is the primary energy source for the brain. Hypoglycemia can cause brain damage or death. Aberrant corticosteroid secretion is responsible for Cushing's and Addison's diseases. Cortisol secretion is controlled primarily by the pituitary peptide ACTH. However, a number of other physiological factors, including Angiotensin II, leptin, nucleotides, epinephrine, and glucose act on AZF cells to regulate secretion of this hormone. While physiological stimuli that regulate cortisol secretion have been identified, the signalling pathways involved are not understood. Although, the release of many hormones is coupled to depolarization-dependent Ca+ entry, the function of specific ion channels in AZF cell secretion has not been clarified. Bovine AZF calls express a novel K+ current (I[AC]) that sets the resting potential while it is inhibited by ACTH, AII, and external ATP at concentrations identical to those that depolarize AZF cells and stimulate cortisol secretion. These channels are also directly activated by intracellular ATP. The Convergent inhibition of I[AC] by three G protein- coupled receptors and its activation by intracellular ATP suggest that this channel is a control point where hormonal and metabolic signals are integrated and transduced to permeability and membrane potential changes associated with cortisol secretion. A detailed characterization of this channel, and the signalling pathways that regulate its activity will be essential to an understanding of adrenal cortical physiology. Whole-cell and single channel patch clamp will be used to describe the modulation of I[AC] K+ channels by hormones and metabolic factors.
The aims of the proposed research will be: (1) to determine whether I[AC] channel gating is coupled to an ATP hydrolysis cycle, allowing these channels to act as glucose sensors; (2) to determine whether other hormonal and paracrine factors which physiologically regulate cortisol secretion, also modulate I[AC] K+ channels; (3) to identify the molecular components of the signalling pathway by which ACTH inhibits I[AC]; and (4) to characterize the properties and distribution of I[AC] K+ channel cDNAs after cloning and expression in a eucaryotic cell line.
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