Measurement and characterization of neuronal cholecystokinin using a novel radioreceptor assay.
This study describes a novel radioreceptor assay (RRA) for cholecystokinin (CCK) which is the first to measure and characterize brain CCK using a technique not dependent on the generation of peptide antibodies. The CCK RRA utilizes the mouse cerebral cortex CCK receptor as the binding source and [125I]BH-CCK-8 as the radiolabelled probe. [125I]BH-CCK-8 bound to the central CCK receptor with a Kd of 1.82 nM and a Bmax of 1.21 pmol/g tissue. Unlabelled CCK-8 displaced the specific binding of [125I]BH-CCK-8 with an inhibition constant of 3.84 nM. CCK was extracted (90% methanol) from discrete brain regions (mouse) and quantified using the CCK RRA. The amygdala contained the highest concentration of CCK (394 +/- 21 pmol/g tissue), followed by the olfactory bulbs (306 +/- 19 pmol/g tissue) and cerebral cortex (298 +/- 21 pmol/g tissue). Moderate levels of CCK were found in the hippocampus (212 +/- 18 pmol/g tissue), striatum (146 +/- 15 pmol/g tissue) and hypothalamus (129 +/- 9 pmol/g tissue). Low levels of CCK were recorded in the pons (45 +/- 5 pmol/g tissue), medulla (41 +/- 3 pmol/g tissue) and spinal cord (29 +/- 3 pmol/g tissue), whilst no CCK was detected in the cerebellum. The molecular forms of CCK in amygdala, cerebral cortex and hypothalamus were characterized using RRA in conjunction with HPLC. CCK-8 was identified as the major molecular form (88%, 94% and 91% of total CCK activity in amygdala, cortex and hypothalamus, respectively) with a smaller component attributable to CCK-4 (8%, 5% and 6% of the total CCK activity).
D000679:Amygdala / Q000032:analysis; D000818:Animals; D001923:Brain Chemistry; D002540:Cerebral Cortex / Q000032:analysis; D002766:Cholecystokinin / Q000032:analysis* / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid; D007031:Hypothalamus / Q000032:analysis; D008297:Male; D051379:Mice; D011869:Radioligand Assay; D012844:Sincalide / Q000378:metabolism
D002766:Cholecystokinin; D012844:Sincalide