Acute and long-term effects of glucose on B-cells from an X-ray induced transplantable rat insulinoma were examined in vitro. Fresh tumour cells failed to respond acutely to glucose or glucose plus theophylline with increased 45Ca uptake and insulin release. During culture in RPMI-1640, blood cells and necrotic tumour cells rapidly disappeared. Cultured secretory cells did not proliferate but exhibited >95% viability and consumed substantial amounts of glucose. Glucose (1.4-22.2 mM) did not affect insulin content or insulin release in 7-day cultures, but both parameters decreased by 51% and 80% compared with 2-day cultures. Insulin output represented 1% of the fall in insulin content suggesting substantial cellular insulin degradation. Acute exposure of 2-day cultured cells to 16.7 mM glucose did not affect insulin release (64-88 ng/106 cells/h) or 45Ca uptake (1.4-2.0 nmol/ 106 cells/h). However, combination with 5 mM theophylline evoked a 22-30% stimulatory insulin response. Cells cultured for 7 days also failed to respond acutely to glucose with increased 45Ca uptake, but a 22-34% stimulation of insulin secretion was evident after maintenance at 5.6-22.2 but not 1.4 mM glucose. Addition of theophylline potentiated the insulin response irrespective of a change in 45Ca uptake.
High animal collagenase (EC 22.214.171.124) activity was detected together with gelatinolytic activity directly in human radicular cyst fluids and in 4 M urea extracts of cyst wall tissues. Both Both cyst wall and cyst fluid collagenases were essentially in latent form and were most effectively activated by p-aminophenylmercuric acetate (APMA). Both APMA-activated collagenases broke down type I collagen preferentially to type III collagen resembling these of polymorphonuclear leukocyte origin.
The influence of extracellular Ca2+ on secretin-induced secretory responses (enzyme output andjuice flow) was examined in the isolated perfused rat pancreas. Secretin in the range of 100pM to 10 nM produced dose-dependent secretory responses. The secretin-induced enzyme output was markedly inhibited when CaCl2 was removed from the perfusing solution, whereas the juice flow by secretin at higher concentrations (1.0 and 10 nM) was not significantly inhibited. Not only the enzyme output but also the juice flow induced by 1 μM forskolin were inhibited in the Ca2+-deficient environment. Continuous stimulation with 100 μM dibutyryl cAMP caused a gradual increase in the pancreatic secretory responses. The enzyme output was abolished but the juice flow was slightly inhibited in the Ca2+-deficient environment. Continuous stimulation with 1 nM secretin induced a gradual increase in the amount of intracellular cyclic AMP, which was significantly decreased in the Ca2+-deficient environment. The present results show that pancreatic enzyme output induced by secretin depends on the extracellular Ca2+-concentration. Two possible mechanisms involved in the Ca2+-dependent enzyme output were discussed in relation to the two kinds of intracellular second messengers, Ca2+ and cyclic AMP.
Toxic principles obtained from the venom of Joro spider (Nephila ciavata) were purified with the reversed phase high performance liquid chromatography and four kinds of the major toxins (JSTX-1, -2, -3 and -4) were isolated. These principles commonly possessed 2,4-dihydroxyphenylacetyl-asparaginyl-cadaveryl moiety in the molecule. The structure of JSTX-3 was determined to be N-(2,4-dihydroxyphenylacetyl-L-asparaginyl)-N'-[N-[4-[(3-aminopropyl)amino]butyl]β-alanyl]cadaverine. The structure activity relationship of the spider toxin was discussed.
Deuterium-labelled p-hydroxyphenylpyruvic acid (PHPPA) was synthesized, and injected intraperitoneally to rats. Deuterium-labelled metabolites excreted into urine were analyzed by means of gas chromatography-negative ion chemical ionization mass spectrometry. During four hours after the injection, the ratio of deuterium-labelled p-hydroxyphenylacetic acid (PHPA) to non-deuterated one reached a high level. On the other hand, the ratio of labelled p-tyramine to non-labelled one remained at a low level. These results indicate that PHPA is biosynthesized not only through deamination of p-tyramine but also through oxidative decarboxylation of PHPPA.
Interactions of retinal projections and VIP-like immunoreactive neurons in the suprachiasmatic nucleus (SCN) of the rat were investigated by horseradish peroxidase (HRP)-histochemical, peptide immunocytochemical, electron microscopic, and morphometric analyses. HRP-filled retinal fibers were detected in the contralateral and ipsilateral part of the ventrolateral part of SCN. Neurons immunoreactive to vasoactive intestinal peptide (VIP), peptide histidine isoleucine amide (PHI), and gastrin releasing peptide (GRP) showed the same distribution as the HRP-filled retinal projections in the intermediate and caudal level of SCN. In immunoelectron microscopy, non-immunoreactive neuropil elements made synaptic contact with VIP-immunoreactive neurons. Degenerating electron microscopic study showed that retinal axon terminals engulfed dendritic spines in SCN. The technique of immunocytochemistry combined with morphometric analysis by color image analyzer revealed that VIP-immunoreactive dots were increased in number 7 days after bilateral enucleation. These results strongly suggest the close interaction between retinal afferents and VIP-containing neurons in SCN.
Histchemical localization of peptide histidine isoleucine (PHI)-like immunoreactivity in the rat brain and spinal cord was investigated using the peroxidase-antiperoxidase technique. PHI-like immunoreactive neurons are principally localized in the cerebral cortex, hippocampus, suprachiasmatic and paraventricular nuclei of the hypothalamus, the central gray and raphe nuclei of the brain stem, aswell as in the spinal nucleus and tract of the trigeminal nerve in the medulla oblongata. PHI-like immunoreactive terminals were densely distributed in the bed nucleus of the stria terminalis, central amygdaloid nucleus, interperduncular nucleus, spinal nucleus and tract of the trigeminal nerve, and dorsal horn of the spinal cord. The origin of PHI-like fibers in the spinal cord and their functional significance are discussed.
The vascular construction of the dog penis was investigated mainly by scanning electron microscopic observation of corrosion casts produced by injection of methylmethacrylate into the penile artery and its branches. The corpus cavernosum penis was supplied mainly by the penile deep artery; only its distal end received a few small branches from the dorsal artery. The helicine arteries were provided with polsters which protruded into the lumen so prominently that they might conspicuously reduce the luminal space during the flaccid state of the penis. The polsters were restricted to the helicine arteries, and could not be found in the penile deep artery or in veins ofthe penis. Cavernous sinuses strikingly anastomosed each other forming a vascular network as a whole. No connection of sinuses was present between the corpus cavernosum penis of either side. Postcavernous venules in the corpus cavernosum penis occurred only on its dorso-medial surface at the point where the crus penis of either side converged; a well-developed capillary network was found in the same region. The postcavernous venules ran for a distance beneath the tunica albuginea, and it is apparent that they are effectively compressed between the tunica albuginea and the peripheral sinuses during erection. In contrast, the cavernous sinuses in the corpus spongiosum urethrae directly emptied into a vein penetrating the thin tunica albuginca. The drainage vein with large calibers was unlikely to receive significant mechanical compression during erection. No arterio-venous anastomoses could be found either within or out of the cavernous bodies.
Cytosolic and cytoskeleton-bound calcium-activated neutral proteases (CANPs) requiring millimolar order Ca2+ were purified from bovine spinal cord, and their action on various neuropeptides and the B chain of oxidized insulin was examined. Both enzymes showed almost the same specificity toward the peptides used as substrates. They cleaved specifically the Arg-Arg bond in dynorphin and the ArgLys bonds in α- and β-neoendorphins, and fairly specifically the Tyr-X and/or PheX bond in neurotensin and angioterisin I. The enzymes also cleaved the Tyr-Leu bond in the B chain of oxidized insulin fairly selectively with additional cleavages occurring to a lesser extent on some other peptide bonds.
In chicken breast muscle myofibrils treated with antiserum against connectin, two distinct stripes were seen in the A-I junction region and less clear stripes in A bands. Anti-nebulin antiserum stained both sides of the Z line, approximately 120 nm apart. Deposits of gold-labeled antibodies against connectin were scattered around the A-I junction region. Gold-labeled antibodies against nebulin were deposited in lines approximately 120 nm apart from the Z line.