The 1989 Upjohn Award Lecture. Cellular and molecular mechanisms in hormone and neurotransmitter secretion

Abstract

Studies on adrenal medulla have had an important influence on the development of a variety of biological concepts, not only within the area of endocrinology, but also in the areas of chemical neurotransmission and secretion in general. The adrenal medulla chromaffin cells are derived embryologically from the neural crest, sharing a common origin with sympathetic neurons and common subcellular features with many endocrine cells. One such feature is the storage of secretory products in membrane-bound organelles, the secretory granules. Secretory cells with these characteristics have been named paraneurons, a term that embraces cells generally and traditionally not considered as neurons, and yet should be regarded as relatives of neurons on the basis of their structure, function, and metabolism. Many of the studies carried out in the past to understand the secretory process have employed perfused adrenal glands. Although this technique has provided very useful information regarding secretion, it did not allow the study of the cellular events involved in the secretory process. To obtain further information on cell secretion, several laboratories including our own have published methods for the isolation and culture of chromaffin cells. The cultured chromaffin cells have shown themselves to be one of the most useful systems developed for the study of the neuroendocrine functions of paraneurons. Studies on cultured chromaffin cells have provided important information on secretory cell cytoskeleton: a group of proteins, some of them previously known from studies on muscle, which form a cytoplasmic network in all non-muscle cells including secretory cells. Immunohistochemical studies have shown at least three types of filament systems: microfilaments, microtubules, and intermediate filaments. In addition, a large variety of cytoskeleton-associated proteins have been characterized. Chromaffin cells are among those non-muscle cells from which cytoskeleton proteins have been isolated and characterized. Owing to similarities between "stimulus–secretion coupling" and "excitation–contraction coupling" in muscle, it has been proposed that the secretory process might be mediated by contractile elements either associated with secretory vesicles or present elsewhere in the secretory cell. Cytoskeletal proteins (actin, myosin, α-actinin, fodrin, tubulin, and neurofilament subunits) and their regulatory proteins (calmodulin, gelsolin) have been isolated from chromaffin cells and characterized. Their physicochemical proteins have been studied and their cellular localizations have been revealed by biochemical, immunocytochemical, and ultrastructural techniques. α-Actinin and fodrin are components of chromaffin granule membranes and some of the cell actin co-purified with secretory granules. Actin forms a network of microfilaments in the subplasmalemma region. This network of filaments is cross-linked and stabilized by several proteins as well as secretory vesicles. Gelsolin, a Ca2+-dependent actin filament severing protein seems to control the length of the actin filaments, thus playing an important role in the regulation of cytoplasm viscosity. Calmodulin also seems to be involved in secretion. Trifluoperazine, a calmodulin antagonist, blocks stimulation-induced hormone release from chromaffin cells at a step distal from calcium entry. High affinity calmodulin binding sites are present in chromaffin granule membranes, and the calmodulin binding proteins of these membranes have been characterized. Furthermore, microinjection of calmodulin antibodies into chromaffin cells blocks hormone output in response to cell stimulation. In view of the above findings, the possible roles of contractile proteins and calmodulin in cell secretion are discussed.Key words: secretion, cytoskeleton, paraneuron, calmodulin, cytosol viscosity.