Showing papers by "Harold Gainer published in 1983"

Dynorphin-A-(1-8) is contained within vasopressin neurosecretory vesicles in rat pituitary

Abstract: Dynorphin-A-(1-8), an opioid peptide widely distributed in the rat central nervous system, is present in vasopressin-containing neurosecretory cells terminating in the neural lobe of the pituitary. Electron microscopic immunocytochemistry reveals that dynorphin-A-(1-8) is contained within the same neurosecretory vesicles as vasopressin and vasopressin-associated neurophysin in the neural lobe of the rat. The results indicate that dynorphin may be released in the pituitary concomitantly with vasopressin during the antidiuretic response.

Optical recording of action potentials from vertebrate nerve terminals using potentiometric probes provides evidence for sodium and calcium components

Abstract: Optical methods are shown to monitor action potentials from a population of nerve terminals in the neurohypophysis of Xenopus. Calcium antagonists such as cadmium and nickel ions block a component of the action potential that probably reflects a calcium-mediated potassium conductance, and tetrodotoxin blocks an inward sodium current, revealing a calcium component to the action potential upstroke.

Precursors of Vasopressin and Oxytocin

Abstract: Publisher Summary It is now known that in addition to transcription and translation processes, various signal sequences and post-translational processing events are also involved in determining the type of peptides that are secreted by the cell. This chapter describes unequivocal criteria for the identification of precursors of peptides with regard to both vasopressin and oxytocin. These criteria include (1) demonstration in classical pulse-chase experiments in intact cellular systems that a larger form of the peptide is first synthesized and subsequently decreases in radioactivity as the radioactive peptide is formed, (2) demonstration by in vitro translation experiments that a larger precursor form of the peptide is synthesized, and (3) utilizing modem recombinant DNA techniques to clone cyclic DNA (cDNA) obtained from a purified messenger RNA (mRNA) template using reverse transcriptase. A variety of biosynthesis studies conducted in vivo and in vitro have shown that the vasopressin precursor is a 19–23,000 molecular weight glycoprotein, which contains vasopressin at its N-terminus, neurophysin in the middle, and a 39 amino acid glycopeptide at the C-terminus. While, the 15–16,000 molecular weight oxytocin precursor is not glycosylated. The vasopressin precursor has been completely sequenced by recombinant DNA methods, thereby providing insights into the nature of the glycopeptide as well as post-translational processing mechanisms. However, many questions remain to be answered such as the nature of the post-translational processing enzymes and mechanisms that fashion the final peptide products or the mechanisms underlying the transcriptional and translational regulation of the precursors during functional activity.

Phospholipid synthesis in the squid giant axon: incorporation of lipid precursors.

Abstract: The squid giant axon and extruded axoplasm from the giant axon were used to study the capacity of axoplasm for phospholipid synthesis. Extruded axoplasm, suspended in chemically defined media, catalyzed the synthesis of phospholipids from all of the precursors tested. 32P-Labeled inorganic phosphate and gamma-labeled ATP were actively incorporated into phosphatidylinositol phosphate, while [2-3H]myo-inositol and L-[3H(G)]serine were actively incorporated into phosphatidylinositol and phosphatidylserine, respectively. Though less well utilized. [2-3H]glycerol was incorporated into phosphatidic acid, phosphatidylinositol, and triglyceride, and methyl-3H]choline and [1-3H]ethanolamine were incorporated into phosphatidylcholine and phosphatidylethanolamine, respectively. Isolated squid giant axons were incubated in artificial seawater containing the above precursors. The axoplasm was extruded following the incubations. Although most of the product lipids were recovered in the sheath (composed of cortical axoplasm, axolemma, and surrounding satellite cells), significant amounts (4-20%) were present in the extruded axoplasm. With tritiated choline and myo-inositol, the major labeled phospholipids found in both the extruded axoplasm and the sheath were phosphatidylcholine and phosphatidylinositol, respectively. With both glycerol and phosphate, phosphatidylethanolamine was a major labeled lipid in both axoplasm and sheath. These findings demonstrate that all classes of phospholipids are formed by endogenous synthetic enzymes in axoplasm. In addition, we feel that the different patterns of incorporation by intact axons and extruded axoplasm indicate that surrounding sheath cells contribute lipids to axoplasm. A comprehensive picture of axonal lipid metabolism should include axoplasmic synthesis and glial-axon transfer as pathways complementing the axonal transport of perikaryally formed lipids.

Calcium-dependent 4-aminopyridine stimulation of protein phosphorylation in squid optic lobe synaptosomes

Abstract: 1. When intact synaptosomes were incubated with [γ-32P]ATP, maximal protein phosphorylation was attained 2 min after the start of incubation. 2. Protein phosphorylation under basal conditions was dependent on external Ca2+, and the dominant peak of phosphorylation was a 50-kd protein. 3. Incubation of intact synaptosomes in the presence of 3–6 mM 4-aminopyridine (4-AP) caused a markedly enhanced phosphorylation of high molecular weight proteins of 90, 100, 130, and 180 kd, with no increase in the 50 or 38 kd proteins. 4. This effect of 4-AP was dependent on external calcium ions in the incubation medium. 5. The 4-AP effect on the high molecular weight proteins was also found in synaptosomal plasma membranes isolated from the synaptosomes. 6. Tetraethylammonium (TEA) ions did not produce this enhancement of phosphorylation.

Vasopressin: an experimental model.

Abstract: The discovery of the Brattleboro rat is a wonderful example of scientific serendipity. In 1961 Henry A. Schroeder, a retired associate professor of clinical physiology at Dartmouth Medical School, noticed that among his private colony of Long-Evans rats housed for research purposes in his summer home in West Brattleboro, Vermont, one cage always had an empty drinking bottle. This cage contained a mother and her litter of 17 pups. Schroeder and his assistant first determined that six of the pups were drinking excessive water and subsequently showed that this behavior could be reversed by treatment with the antidiuretic hormone, vasopressin. The Brattleboro rat was thus born, and the pups were then nurtured by Valtin and Sokol to become the Brattleboro strain at Dartmouth Medical School. The discovery of the rat is described by Valtin in the first paper in this volume, a compilation of 43 invited papers and 50 shorter research reports that were presented at an international symposium celebrating the 20th anniversary of the Brattleboro rat. About 120 scientists from four continents gathered to discuss the total biology of this mutant rat strain. The principal interest in the Brattleboro rat is that it is unable to synthesize vasopressin and hence is valuable as an experimental model for diabetes insipidus. The mode of inheritance of the genetic defect is semirecessive (that is, the heterozygote has a vasopressin content intermediate between the homozygote and normals of the Long-Evans parent strain). Descendants of the original strain developed by Valtin and Sokol are now widely distributed and bred in laboratories all over the world. One of the major issues this book discusses is that these different laboratories have introduced the gene into diverse inbred rat stocks, thereby producing vasopressindeficient rats with heterogeneous genetic backgrounds. This may be the cause of a number of discrepancies in the findings of different laboratories using the \"Brattleboro rat,\" especially in studies that deal with highly complex phenomena related to vasopressin (such as behavioral studies). Therefore, the first part of