sq-23377 and Acidosis

Previous studies concerning ischemic priapism revealed that hypoxia alters the erectile and contractile responses of penis. But the effects of accompanying acidosis on those responses have not been fully evaluated or understood yet. We performed this study to elucidate the role of acidosis on the trabecular smooth muscle contractility like in ischemic priapism. Under the general anesthesia, 55 mature male cats were conditioned to systemic metabolic acidosis by hypoventilation by animal ventilator. The changes of intracavernous pressure (ICP) to erectogenic agents (acetylcholine, L-arginine, prostaglandin E1: PGE1), erectolytic agents (epinephrine, thromboxane A2; TXA2), K channel-related drugs (pinacidil, 4-aminopyridine, tetraethylammonium; TEA, glibenclamide) and calcium ionophore were monitored at Set 1 (PO2 > 60 mmHg, pH > 7.25), Set 2 (PO2 < 30 mmHg, 7.25 > pH > 7.0), Set 3 (PO2 < 30 mmHg, pH < 7.0), and Set 4 (PO2 > 60 mmHg, pH < 7.0) in vivo. At Set 1 and Set 2, epinephrine, TXA2, and ionomycin decreased the ICP by acetylcholine or PGE1 (n = 9, P < 0.01). The decrease of ICP was in order of epinephrine, TXA2 and ionomycin. Acidosis reduced the increase of ICP to acetylcholine or PGE1 (n = 8, P < 0.01), TXA2 or ionomycin did not affect ICP under severe acidosis but epinephrine decreased ICP even under severe acidosis (n = 7, P < 0.05). Pretreatment of potassium channel blockers did not suppress the increase of ICP by erectogenic agents under acidosis (n = 6, P < 0.05). Pinacidil did not affect ICP under acidosis (n = 6, P < 0.01). These results suggest that acidosis impairs the contractile response of cavernous smooth muscle to erectolytic agents. It may be the results of the interference by [H+] with the intra and extracellular mechanisms that regulate the homeostasis of [Ca2]. Conclusively, besides hypoxia, acidosis is another limiting factor of trabecular smooth muscle contractility like in ischemic priapism.

Topics: Acetylcholine; Acidosis; Alprostadil; Animals; Arginine; Cats; Epinephrine; Hypoxia; Ionomycin; Male; Muscle Contraction; Muscle, Smooth; Penile Erection; Penis; Potassium Channel Blockers; Pressure; Thromboxane A2

We have examined the dependence of unesterified fatty acid accumulation by intact, freshly isolated proximal tubules on Ca2+, pH, and the cytoprotective amino acid, glycine, during injury induced by hypoxia, antimycin, or antimycin plus ionomycin. In the absence of glycine, similarly high levels of fatty acid accumulation were seen during all three injury conditions irrespective of whether tubules were incubated in normal 1.25 mM Ca2+ medium or in medium where Ca2+ was buffered to 0.1 microM, a maneuver which prevented injury-associated increase of cytosolic-free Ca2+ as measured with fura 2. In the presence of glycine, which strongly suppressed development of lethal membrane damage for at least 60 min and did not have any apparent direct effects on fatty acid accumulation, both Ca(2+)-independent and Ca(2+)-dependent components of fatty acid accumulation were discernible. The Ca(2+)-independent component accounted for approximately 2/3 of fatty acid accumulation and did not vary as Ca2+ ranged from 10 nM to 1 microM. Unequivocal Ca(2+)-dependent accumulation occurred when Ca2+ exceeded 10 microM. Lowering pH to 6.9 had a moderate, generalized suppressive effect on fatty acid accumulation, including the major Ca(2+)-independent component, irrespective of the presence of glycine. These data emphasize the role of Ca(2+)-independent fatty acid accumulation during proximal tubule cell injury, clarify the modulatory actions of the potent, intrinsic cytoprotective factors, glycine and reduced pH, and provide insight into the relationship between fatty acid accumulation and lethal membrane damage.

Topics: Acidosis; Animals; Antimycin A; Biological Transport; Calcium; Cell Hypoxia; Dose-Response Relationship, Drug; Drug Interactions; Egtazic Acid; Fatty Acids, Nonesterified; Glycine; Hydrogen-Ion Concentration; In Vitro Techniques; Ionomycin; Kidney Cortex; Kidney Tubules, Proximal; Kinetics; L-Lactate Dehydrogenase; Male; Phospholipids; Rabbits; Time Factors

These studies examined the effect of acidosis on immediate early (IE) gene expression in renal tubule cells. In MCT cells, an SV40 transformed mouse proximal tubule cell line, incubation in acid media led to transient increases in c-fos, c-jun, junB, and egr-1 mRNA abundance, peaking at 30 min to 1 h. In vivo metabolic acidosis caused more prolonged increases in these mRNA species in renal cortex. Nuclear runon studies demonstrated increased rates of transcription for these IE genes. In addition, pretreatment of cells with cycloheximide caused superinduction of these mRNA by acid incubation. These responses are similar to those elicited by growth factors. Inhibition of tyrosine kinase pathways prevented IE gene activation by acid, while inhibition of protein kinase C and/or increases in cell calcium had no effect. In 3T3 cells, acid activated IE genes by a different mechanism in that the increase in mRNA did not include c-jun, was more prolonged, and was blocked by cycloheximide. In summary, incubation of renal cells in acid media leads to activation of IE genes that is similar to growth factor-induced IE gene activation, and is likely mediated by tyrosine kinase pathways.

Topics: 3T3 Cells; Acidosis; Animals; Cell Line; Cell Nucleus; DNA-Binding Proteins; Early Growth Response Protein 1; Epithelium; Gene Expression Regulation; Genes, Immediate-Early; Hydrogen-Ion Concentration; Immediate-Early Proteins; Ionomycin; Kidney Tubules, Proximal; Mice; Proto-Oncogene Proteins c-fos; Proto-Oncogene Proteins c-jun; RNA, Messenger; Tetradecanoylphorbol Acetate; Transcription Factors; Transcriptional Activation

To assess the role of increased cytosolic free calcium (Caf) in the pathogenesis of acute proximal tubule cell injury and the protection afforded by exposure to reduced medium pH or treatment with glycine, fura-2-loaded tubules were studied in suspension and singly in a superfusion system. The Ca2+ ionophore, ionomycin, increased Caf to micromolar levels and rapidly produced lethal cell injury as indicated by loss of lactate dehydrogenase to the medium by suspended tubules and accelerated leak of fura and failure to exclude Trypan blue by superfused tubules. Decreasing medium Ca2+ to 100 nM prevented the ionomycin-induced increases of Caf and the injury. Reducing medium pH from 7.4 to 6.9 or adding 2 mM glycine to the medium also prevented the cell death, but did not prevent the increase of Caf to micromolar levels. Cells treated with 1799, an uncoupler of oxidative phosphorylation which produced severe adenosine triphosphate (ATP) depletion, did not develop increases of Caf until just before loss of viability. Preventing these increases of Caf with 100 nM Ca2+ medium did not protect 1799-treated cells. Reduced pH and glycine protected 1799-treated cells without ameliorating the increases of Caf. These data demonstrate the toxic potential of increased Caf in the proximal tubule and show that Caf does sharply increase prior to loss of viability in an ATP depletion model of injury, but this increase does not necessarily contribute to the outcome. The potent protective actions of decreased pH and glycine allow the cells to sustain increases of Caf to micromolar levels in spite of severe, accompanying cellular ATP depletion without developing lethal cell injury.

Topics: Acidosis; Adenosine Triphosphate; Animals; Calcium; Cell Survival; Cytosol; Fura-2; Glycine; Hydrogen-Ion Concentration; Ionomycin; Kidney Tubules, Proximal; L-Lactate Dehydrogenase; Male; Rabbits