Glomerular size-selective dysfunction in NIDDM is not ameliorated by ACE inhibition or by calcium channel blockade.
BACKGROUND: In patients with insulin-dependent diabetes mellitus (IDDM) and overt nephropathy glomerular barrier size-selectivity progressively deteriorates with time and is effectively improved by angiotensin converting enzyme (ACE) inhibition. Whether similar glomerular functional changes develop in proteinuric patients with non-insulin-dependent diabetes mellitus (NIDDM), and whether antihypertensive agents can favorably affect glomerular filtration of macromolecules in these patients, has not been documented yet. METHODS: We investigated renal hemodynamics and fractional clearance of neutral dextrans of graded sizes, in nine proteinuric patients with NIDDM and renal biopsy findings of typical diabetic glomerulopathy. Six healthy volunteers served as controls. We also investigated the effects of an ACE inhibitor and of a calcium channel blocker, both given in doses targeted to achieve a comparable level of systemic blood pressure control, on glomerular hemodynamics and sieving function. Theoretical analysis of glomerular macromolecule transport was adopted to evaluate intrinsic glomerular membrane permeability properties. RESULTS: Fractional clearance of large macromolecules (42 to 66 A in radius) was significantly higher in diabetic patients than in controls, and the distribution of membrane pore radii was calculated to be shifted towards larger pore sizes in diabetics (mean radius increased from 55 to 60 A). Despite effective blood pressure control, neither antihypertensive affected glomerular hemodynamics to any significant extent. Fractional clearance of dextrans, as well as of albumin and IgG, and total urinary proteins were not modified by either treatments. CONCLUSIONS: These data indicate that patients with NIDDM and overt nephropathy develop abnormalities in size-selective function of the glomerular barrier and, at variance to IDDM, such changes were not ameliorated either by ACE inhibition or calcium channel blockade. (+info)
Retinal input induces three firing patterns in neurons of the superficial superior colliculus of neonatal rats.
By using an in vitro isolated brain stem preparation, we recorded extracellular responses to electrical stimulation of the optic tract (OT) from 71 neurons in the superficial superior colliculus (SC) of neonatal rats (P1-13). At postnatal day 1 (P1), all tested neurons (n = 10) already received excitatory input from the retina. Sixty-nine (97%) superficial SC neurons of neonatal rats showed three response patterns to OT stimulation, which depended on stimulus intensity. A weak stimulus evoked only one spike that was caused by activation of non-N-methyl-D-aspartate (NMDA) glutamate receptors. A moderate stimulus elicited a short train (<250 ms) of spikes, which was induced by activation of both NMDA and non-NMDA receptors. A strong stimulus gave rise to a long train (>300 ms) of spikes, which was associated with additional activation of L-type high-threshold calcium channels. The long train firing pattern could also be induced either by temporal summation of retinal inputs or by blocking gamma-aminobutyric acid-A receptors. Because retinal ganglion cells show synchronous bursting activity before eye opening at P14, the retinotectal inputs appear to be sufficient to activate L-type calcium channels in the absence of pattern vision. Therefore activation of L-type calcium channels is likely to be an important source for calcium influx into SC neurons in neonatal rats. (+info)
NMDA receptor characterization and subunit expression in rat cultured mesencephalic neurones.
1. NMDA-induced changes in free intracellular Ca2+ concentration ([Ca2+]i) were determined in individual cultured rat mesencephalic neurones by the fura-2 method. mRNA expression encoding NMDA receptor subunits (NR1, NR2A-D) was examined by RT-PCR. 2. NMDA (1-100 microM, plus 10 microM glycine) induced a concentration-dependent increase in [Ca2+]i (EC50 = 5.7 microM). The effect of NMDA was virtually insensitive to tetrodotoxin (0.3 microM) and nitrendipine (1 microM), but dependent on extracellular Ca2+. 5,7-Dichlorokynurenic acid (10 microM), a specific antagonist at the glycine binding site on the NMDA receptor, abolished the NMDA response. 3. Memantine, an open-channel blocker, and ifenprodil, a preferential non-competitive NR1/NR2B receptor antagonist diminished the NMDA effect with an IC50 value of 0.17 and 1 microM, respectively. Ethanol at 50 and 100 mM caused about 25 and 45%-inhibition, respectively. 4. Agarose gel analysis of the PCR products followed by ethidium bromide fluorescence or CSPD chemiluminescence detection revealed an almost exclusive expression of the NR1 splice variants lacking exon (E) 5 and E22. The 3' splice form without both E21 and E22 exceeded that containing E21 by approximately 4 fold. The relative amounts of NR2A, NR2B, NR2C corresponded to approximately 1:2:1. NR2D mRNA was also detectable. 5. In conclusion, mesencephalic neurones bear ethanol-sensitive NMDA receptors which might be involved in the development of ethanol dependence and withdrawal. The high affinity of NMDA to this receptor, its sensitivity to ifenprodil and memantine may suggest that the mesencephalic NMDA receptor comprises the NR1 splice variant lacking E5, NR2B, and NR2C, respectively. (+info)
Differences in the actions of some blockers of the calcium-activated potassium permeability in mammalian red cells.
1. The actions of some inhibitors of the Ca2+-activated K+ permeability in mammalian red cells have been compared. 2. Block of the permeability was assessed from the reduction in the net loss of K+ that followed the application of the Ca2+ ionophore A23187 (2 microM) to rabbit red cells suspended at a haematocrit of 1% in a low potassium solution ([K]0 0.12-0.17 mM) at 37 degrees C. Net movement of K+ was measured using a K+-sensitive electrode placed in the suspension. 3. The concentrations (microM +/- s.d.) of the compounds tested causing 50% inhibition of K+ loss were: quinine, 37 +/- 3; cetiedil, 26 +/- 1; the cetiedil congeners UCL 1269, UCL 1274 and UCL 1495, approximately 150, 8.2 +/- 0.1, 0.92 +/- 0.03 respectively; clotrimazole, 1.2 +/- 0.1; nitrendipine, 3.6 +/- 0.5 and charybdotoxin, 0.015 +/- 0.002. 4. The characteristics of the block suggested that compounds could be placed in two groups. For one set (quinine, cetiedil, and the UCL congeners), the concentration-inhibition curves were steeper (Hill coefficient, nH, > or = 2.7) than for the other (clotrimazole, nitrendipine, charybdotoxin) for which nH approximately 1. 5. Compounds in the first set alone became less active on raising the concentration of K+ in the external solution to 5.4 mM. 6. The rate of K+ loss induced by A23187 slowed in the presence of high concentrations of cetiedil and its analogues, suggesting a use-dependent component to the inhibitory action. This was not seen with clotrimazole. 7. The blocking action of the cetiedil analogue UCL 1274 could not be overcome by an increase in external Ca2+ and its potency was unaltered when K+ loss was induced by the application of Pb2+ (10 microM) rather than by A23187. 8. These results, taken with the findings of others, suggest that agents that block the red cell Ca2+-activated K+ permeability can be placed in two groups with different mechanisms of action. The differences can be explained by supposing that clotrimazole and charybdotoxin act at the outer face of the channel whereas cetiedil and its congeners may block within it, either at or near the K+ binding site that determines the flow of K+. (+info)
Effects of calcium-channel blockade in older patients with diabetes and systolic hypertension. Systolic Hypertension in Europe Trial Investigators.
BACKGROUND: Recent reports suggest that calcium-channel blockers may be harmful in patients with diabetes and hypertension. We previously reported that antihypertensive treatment with the calcium-channel blocker nitrendipine reduced the risk of cardiovascular events. In this post hoc analysis, we compared the outcome of treatment with nitrendipine in diabetic and nondiabetic patients. METHODS: After stratification according to center, sex, and presence or absence of previous cardiovascular complications, 4695 patients (age, > or =60 years) with systolic blood pressure of 160 to 219 mm Hg and diastolic pressure below 95 mm Hg were randomly assigned to receive active treatment or placebo. Active treatment consisted of nitrendipine (10 to 40 mg per day) with the possible addition or substitution of enalapril (5 to 20 mg per day) or hydrochlorothiazide (12.5 to 25 mg per day) or both, titrated to reduce the systolic blood pressure by at least 20 mm Hg and to less than 150 mm Hg. In the control group, matching placebo tablets were administered similarly. RESULTS: At randomization, 492 patients (10.5 percent) had diabetes. After a median follow-up of two years, the systolic and diastolic blood pressures in the placebo and active-treatment groups differed by 8.6 and 3.9 mm Hg, respectively, among the diabetic patients. Among the 4203 patients without diabetes, systolic and diastolic pressures differed by 10.3 and 4.5 mm Hg, respectively, in the two groups. After adjustment for possible confounders, active treatment was found to have reduced overall mortality by 55 percent (from 45.1 deaths per 1000 patients to 26.4 deaths per 1000 patients), mortality from cardiovascular disease by 76 percent, all cardiovascular events combined by 69 percent, fatal and nonfatal strokes by 73 percent, and all cardiac events combined by 63 percent in the group of patients with diabetes. Among the nondiabetic patients, active treatment decreased all cardiovascular events combined by 26 percent and fatal and nonfatal strokes by 38 percent. In the group of patients receiving active treatment, reductions in overall mortality, mortality from cardiovascular disease, and all cardiovascular events were significantly larger among the diabetic patients than among the nondiabetic patients (P=0.04, P=0.02, and P=0.01, respectively). CONCLUSIONS: Nitrendipine-based antihypertensive therapy is particularly beneficial in older patients with diabetes and isolated systolic hypertension. Thus, our findings do not support the hypothesis that the use of long-acting calcium-channel blockers may be harmful in diabetic patients. (+info)
Correlation between increases in dihydropyridine binding in vivo and behavioural signs of ethanol withdrawal in mice.
Increased ligand binding to dihydropyridine receptors in the central nervous system after chronic ethanol consumption is thought to contribute to the withdrawal syndrome. Previous studies demonstrated such changes when the binding was measured in vitro, which, as the receptors are voltage-sensitive, may not accurately reflect the binding in vivo. In the present study, dihydropyridine binding was measured in vivo in mice, after intravenous administration of the radioligand. The aim was to determine whether there was any correlation between such binding and measurements of behavioural hyperexcitability at different times during the withdrawal phase and after two different methods of alcohol administration. Measurements were made of the binding in vivo of [3H]nitrendipine, at intervals after withdrawal from chronic ethanol administration, and of the severity of withdrawal as measured by response to gentle handling. An increase in the in vivo binding to [3H]nitrendipine was seen after cessation of chronic ethanol consumption by liquid diet. The binding was significantly increased at 4 h, when the behavioural changes were maximal, but not immediately after withdrawal, when the responses to handling were unchanged. By 24 h after cessation of the ethanol treatment, no differences in the binding were found, compared with control values; at this time the withdrawal hyperexcitability had ceased. When alcohol was given chronically by inhalation, the in vivo dihydropyridine binding was increased at 3 h from withdrawal of the ethanol, the time of maximal behavioural hyperexcitability, but no change was seen 30 min after withdrawal, when no changes in the ratings of behaviour were found. There was a significant positive correlation in individual mice between the ratings of handling-induced behaviour at the 3 h interval and the amount of in vivo binding. These data support the hypothesis that the sites labelled by [3H]nitrendipine play an important role in withdrawal hyperexcitability. (+info)
Intracellular Ca2+ concentrations in cultured chicken photoreceptor cells: sustained elevation in depolarized cells and the role of dihydropyridine-sensitive Ca2+ channels.
PURPOSE: Retinal photoreceptor cells are tonically depolarized in darkness. Ca2+ influx in darkness plays a critical role in the regulation of neurotransmitter release and melatonin synthesis in these sensory cells. The purpose of the present study was to examine the dynamic changes of intracellular Ca2+ concentrations ([Ca2+]in ) in response to a tonic depolarizing stimulus and to determine the role of dihydropyridine-sensitive calcium channels in the response. METHODS: Photoreceptor cells were prepared from embryonic chick retina and cultured for 6-12 days. Cells were depolarized by exposure to 35 mM extracellular K+. [Ca2+]in of individual photoreceptor cell bodies/synaptic terminals was determined by ratiometric fura-2 image analysis. RESULTS: Chemical depolarization with 35 mM [K+]out greatly increased [Ca2+]in of inner segment/synaptic terminal regions of photoreceptors. The increase usually reached a plateau after the first few minutes of stimulation and was sustained for prolonged periods (>2 h) in the presence of high K+. When the extracellular K+ concentration was reduced, the [Ca2+]in rapidly returned to the basal level. Substitution of 1 mM CoCl2 for CaCl2 in the superfusion medium rapidly and reversibly reduced the [Ca2+]in of depolarized photoreceptor cells. Antagonists of L-type Ca2+ channels, nitrendipine and nifedipine, inhibited the K+-evoked increase of [Ca2+]in. Bay K 8644, a dihydropyridine Ca2+ channel agonist, potentiated the increase of [Ca2+]in elicited by high K+. In some cells, Bay K 8644 alone increased [Ca2+]in under basal conditions. CONCLUSIONS: The increase of [Ca2+]in elicited by depolarization with 35 mM extracellular K+ is due to influx of calcium through the dihydropyridine-sensitive voltage-gated channels. Intracellular [Ca2+] remains elevated for extended periods of time during tonic depolarization. This sustained response requires continuous Ca2+ channel activity. (+info)
Genistein inhibits pressure-induced expression of c-fos in isolated mesenteric arteries.
We have previously demonstrated that elevating intraluminal pressure from 90 to 140 mm Hg in isolated mesenteric arteries increases the expression of proto-oncogenes. These proto-oncogenes encode nuclear transcription factors that regulate the expression of target genes during various stages of the cell cycle. Thus, pressure-induced proto-oncogene expression may represent a mechanism by which pressure can induce growth and/or proliferation of vascular smooth muscle. The purpose of this study was to determine the intracellular signals that contribute to the pressure-induced increase in c-fos expression. Small mesenteric arteries were isolated from male Wistar rats and transferred to a dual-vessel chamber. The arteries were cannulated and slowly equilibrated to initial conditions (90 mm Hg, 37 degrees C) while being continuously superfused with a HEPES-bicarbonate-buffered Krebs' solution. After the equilibration period, the intraluminal pressure in 1 artery was increased to 140 mm Hg for 1 hour. In experiments designed to determine the intracellular signals involved in the pressure-induced increase in c-fos expression, specific inhibitors were introduced to the superfusate reservoir of both arteries before the pressure increase. The arteries were then fixed in phosphate-buffered formalin and embedded in paraffin blocks. Sections of paraffin-embedded arteries were fixed on slides, and the expression of c-fos was determined by in situ hybridization with the use of (35)S-labeled riboprobes. The pressure-induced expression of c-fos was not inhibited by nitrendipine (10 micromol/L), a calcium-free Krebs' solution containing EGTA (1 to 2 mmol/L), calphostin C (0.1 micromol/L), or cytochalasin D (0.4 micromol/L) but was inhibited by genistein (30 micromol/L). The results suggest that activation of a tyrosine kinase is required for pressure-induced c-fos expression, but the signaling pathway does not require extracellular calcium entry, intact actin filaments, or protein kinase C. As we have shown previously, the expression of c-fos correlated with wall stress. (+info)