Sympathetic axons and nerve terminals: the protein composition of small and large dense-core and of a third type of vesicles

Abstract

Homogenates of bovine splenic nerve and of vas deferens were subjected to differential and density gradient centrifugation to investigate their noradrenaline-storing organelles. The subcellular fractions obtained were analysed by immunoblotting in order to define the presence of various antigens in small dense-core and large dense-core vesicles. In both large granule and microsomal fractions from splenic nerve only one type of noradrenaline-storing vesicle was found, which represents the large dense-core vesicles. These organelles contained chromogranin A, chromogranin B, cytochrome b-561, carboxypeptidase H, glycoprotein II, glycoprotein III, dopamine beta-hydroxylase and the monoamine carrier which are also present in adrenal chromaffin granules. The subcellular distribution of synaptin/synatophysin was more complex since this protein was apparently present in two organelles: in a light vesicle which did not contain significant amounts of antigens found in large dense-core vesicles (dopamine beta-hydroxylase, cytochrome b-561 and the monoamine carrier) and in the dense fractions of the gradient, possibly within large dense-core vesicles. In the microsomal gradient from vas deferens several markers (catecholamines, synaptin/synaptophysin and dopamine beta-hydroxylase) were found in a bimodal distribution, which is consistent with their presence in small and large dense-core vesicles. When the larger granules were removed with higher centrifugation speed a microsomal fraction containing only light vesicles was obtained. After gradient centrifugation of this fraction several components (catecholamines, dopamine beta-hydroxylase, cytochrome b-561, the monoamine carrier and synaptin/synaptophysin) were concentrated in a peak at low density; apparently only small dense-core vesicles were now present.(ABSTRACT TRUNCATED AT 250 WORDS)