Chromogranin B gene ablation reduces the catecholamine cargo and decelerates exocytosis in chromaffin secretory vesicles.

Abstract

Chromogranins/secretogranins (Cgs) are the major soluble proteins of large dense-core secretory vesicles (LDCVs). We have recently reported that the absence of chromogranin A (CgA) caused important changes in the accumulation and in the exocytosis of catecholamines (CAs) using a CgA-knock-out (CgA-KO) mouse. Here, we have analyzed a CgB-KO mouse strain that can be maintained in homozygosis. These mice have 36% less adrenomedullary epinephrine when compared to Chgb / [wild type (WT)], whereas the norepinephrine content was similar. The total evoked release of CA was 33% lower than WT mice. This decrease was not due to a lower frequency of exocytotic events but to less secretion per quantum ( 30%) measured by amperometry; amperometric spikes exhibited a slowerascendingbutanormaldecayingphase.CellincubationwithL-DOPAincreasedthevesicleCAcontentofWTbutnotoftheCgB-KO cells. Intracellular electrochemistry, using patchamperometry,showedthatL-DOPAoverloadproducedasignificantlylargerincreasein cytosolic CAsincellsfromtheKOanimalsthanchromaffincellsfromtheWT.Thesedataindicatethatthemechanismsforvesicular accumulation of CAs in the CgB-KO cells were saturated, while there was ample capacity for further accumulation in WT cells. Protein analysis of LDCVs showed the overexpression of CgA as well as other proteins apparently unrelated to the secretory process. We conclude that CgB, like CgA, is a highly efficient system directly involved in monoamine accumulation and in the kinetics of exocytosis from LDCVs.