Measuring of extracellular cortisol and corticotropin-releasing hormone inthe amygdala using immunosensor coupled microdialysis

A method is described for the measurement of cortisol or corticotropin-rele asing hormone (CRH) from the brain, within, or in the dialysate of, a micro dialysis probe using an antibody-linked assay. Polyclonal antibodies for ei ther cortisol or CRH provide a specificity of measurement. These antibodies are affixed on a platinum electrode within the probe. Determination of bou nd cortisol or CRH is performed via an indirect assessment of competitive l igand also bound, and conjugated to HRP. This is activated and measured as current change. These probes were used for extracellular measurements in th e amygdala, a limbic brain region, of sheep. Direct measurement in-vivo com pared favorably to the use of either the immunosensor or standard RIA techn iques on dialysate emerging from a classical microdialysis probe, suggestin g that the method could be used directly in-vivo. This is the first report in literature of rapid on-line measurement of CRH and cortisol from a discr ete brain area. The probes have fast response times ( > 90% of maximum resp onse within 30 s of start of analysis), high sensitivity ( < 0.1 pg/ml for CRH) and can acquire data every 2 min. They are stable in-vivo ( > 72 h) an d regenerable, offering on-line measurement, with rapid time resolution, of neurohumoral substances. (C) 2001 Elsevier Science BN. All rights reserved .