Effects of Ca(2+) channel activity on renal hemodynamics during acute attenuation of NO synthesis in the rat.

In cultured vascular muscle cells, nitric oxide (NO) has been shown to inhibit voltage-dependent Ca(2+) channels, which are involved in renal blood flow (RBF) autoregulation. Therefore, our purpose was to specify in vivo the effects of this interaction on RBF autoregulation. To do so, hemodynamics were investigated in anesthetized rats during Ca(2+) channel blockade before or after acute NO synthesis inhibition. Rats were treated intravenously with vehicle (n = 10), 0.3 mg/kg body wt N(G)-nitro-L-arginine-methyl ester (L-NAME; n = 7), 4.5 microg. kg body wt(-1). min(-1) nifedipine (n = 8) alone, or with nifedipine infused before (n = 8), after (n = 8), or coadministered with L-NAME (n = 10). Baseline renal vascular resistance (RVR) averaged 14.0 +/- 1.2 resistance units and did not change after vehicle. RVR increased or decreased significantly by 27 and 29% after L-NAME or nifedipine, respectively. Nifedipine reversed, but did not prevent, RVR increase after or coadministered with L-NAME. RBF autoregulation was maintained after L-NAME, but the autoregulatory pressure limit (P(A)) was significantly lowered by 15 mmHg. Nifedipine pretreatment or coadministration with L-NAME limited P(A) resetting or suppressed autoregulation at higher doses. Results were similar with verapamil. Intrarenal blockade of Ca(2+)-activated K(+) channels also prevented autoregulatory resetting by L-NAME (n = 8). These findings suggest NO inhibits voltage-dependent Ca(2+) channels and thereby modulates RBF autoregulatory efficiency.

Lysosomal alterations induced in cultured rat fibroblasts by long-term exposure to low concentrations of azithromycin.

Computer-aided simulations suggest that the doses and schedules of administration of azithromycin proposed in treatment and prophylaxis of Mycobacterium avium complex (MAC) in AIDS patients will result in drug concentrations in serum and extracellular fluids remaining for sustained periods of time in the 0.03-0.1 mg/L range. We exposed cultured rat embryo fibroblasts to these concentrations (and multiples up to 20 mg/L) for up to 16 days. Electron microscopy showed that after 7 days' incubation in 0.03 mg/L azithromycin, there was conspicuous accumulation of osmiophilic, lamellar structures (myeloid bodies) in lysosomes, suggesting the onset of a phospholipidosis. Assay of total cell phospholipids and cholesterol showed significant increases in cells exposed to > or = 1 to 5 mg/L of azithromycin in association with hyperactivity of the lysosomal enzyme cathepsin B. The data suggest that azithromycin, at extracellular concentrations pertinent to its use for MAC treatment, and perhaps also prophylaxis, causes limited morphological alterations of the lysosomes in cultured cells which are of the same nature as those developing rapidly and extensively at higher concentrations.

Hyperactivity of cathepsin B and other lysosomal enzymes in fibroblasts exposed to azithromycin, a dicationic macrolide antibiotic with exceptional tissue accumulation.

Azithromycin accumulates in lysosomes where it causes phospholipidosis. In homogenates prepared by sonication of fibroblasts incubated for 3 days with azithromycin (66 microM), the activities of sulfatase A, phospholipase A1, N-acetyl-beta-hexosaminidase and cathepsin B increased from 180 to 330%, but not those of 3 non-lysosomal enzymes. The level of cathepsin B mRNA was unaffected. The hyperactivity induced by azithromycin is non-reversible upon drug withdrawal, prevented by coincubation with cycloheximide, affects the Vmax but not the Km, and is not reproduced with gentamicin, another drug also causing lysosomal phospholipidosis. The data therefore suggest that azithromycin increases the level of lysosomal enzymes by a mechanism distinct from the stimulation of gene expression but requiring protein synthesis, and is not in direct relation to the lysosomal phospholipidosis.