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Beschreibung

Millipore's Ready-to-Assay GPCR frozen cells are designed for simple, rapid calcium assays with no requirement for intensive cell culturing. Millipore has optimized the freezing conditions to provide cells with high viability and functionality post-thaw. The user simply thaws the cells and resuspends them in media, dispenses cell suspension into assay plates and, following over night recovery, assays for calcium response. The endogenous catecholamines epinephrine and norepinephrine have profound effects on smooth muscle activity, cardiac function, carbohydrate and fat metabolism, hormone secretion, neurotransmitter release, and central nervous system actions. These activities are mediated by GPCRs belonging to two subfamilies, the alpha- and beta-adrenoceptors (Bylund et al., 1994). The alpha2 adrenergic receptor subfamily members, consisting of alpha2A, alpha2B, and alpha2C, couple primarily to Gi to inhibit cAMP production, and play an important role in regulation of cardiovascular and CNS function. The alpha2A receptor at presynaptic sites has an inhibitory effect on catecholamine release from sympathetic nerve endings. Experiments with alpha2A -selective agonists and mice lacking alpha2A demonstrate that activation of alpha2A results in hypotension, sedation, analgesia, and hypothermia (Kable et al., 2000). Millipore's cloned human alpha2A -expressing cell line is made in the Chem-1 host, which supports high levels of recombinant alpha2A expression on the cell surface and contains high levels of the promiscuous G protein Galpha15 to couple the receptor to the calcium signaling pathway. Thus, the cell line is an ideal tool for screening for agonists, antagonists and modulators at alpha2A.