Induction of manganese superoxide dismutase in BV-2 microglial cells.

The regulation of the manganese-dependent superoxide dismutase (Mn-SOD) was studied in immortalized microglial cells (line BV-2). BV-2 cells, activated with lipopolysaccharide (LPS), exhibited an increase in Mn-SOD-like immunoreactivity, that was associated with an accumulation of nitrite in the culture medium and an increase in immunoreactivity for the inducible type of nitric oxide synthase (i-NOS). The i-NOS inhibitor L-N6-(1-iminoethyl)-lysine (NIL, 600 microM) suppressed the nitrite accumulation and the increase in Mn-SOD-like immunoreactivity in activated cells without significant effect on the level of i-NOS-like immunoreactivity. The NO donor sodium nitroprusside dose-dependently increased Mn-SOD-like immunoreactivity in NIL-pretreated BV-2 cells. These results indicate that the induction of Mn-SOD in activated BV-2 cells is mediated in part by NO, or its metabolites.

Hippocampal muscarinic receptor function in spatial learning-impaired aged rats.

Efficiency of coupling of hippocampal muscarinic receptors to phosphoinositide (PI) turnover was investigated in behaviorally characterized young and aged Long-Evans rats using hippocampal minces and the method of partial receptor alkylation of Furchgott. Densities of the m1, m2, and m3 receptor proteins were determined using specific antibodies and immunoprecipitation. Spatial learning ability was quantified using a water maze. There were no differences in the levels of muscarinic receptor proteins between young and aged (27 months) rats or in rats with impaired spatial learning. The dissociation constant (KD) for the agonist oxotremorine-M and the KD/EC50 ratio, an indicator of receptor-effector coupling efficiency were similar in young and aged rats. However, the maximal PI turnover response to oxotremorine-M was decreased in impaired aged rats and this parameter was highly correlated with the spatial learning index (R = -0.825; p < 0.001). A reduction in effector stimulation in the absence of changes in receptor protein or coupling efficiency suggests that dysfunction in the hippocampal muscarinic receptor systems occurs at the level of phospholipase C or beyond.

Presence of the N-methyl-D-aspartate-associated glycine receptor agonist, D-serine, in human temporal cortex: comparison of normal, Parkinson, and Alzheimer tissues.

The amino acid D-serine (D-Ser), previously recognized as a pharmacological tool for potentiating neuronal activity mediated by the N-methyl-D-aspartate (NMDA) receptor complex, in vitro and in vivo, has been observed in several brain regions of the rat and mouse, most prominently in cortex. In addition to reconfirming the presence and distribution of D-Ser in rat brain, we have observed, for the first time, endogenous, free D-Ser in temporal cortex of normal human brains at a level of 2.18 +/- 0.12 nmol/mg of protein, representing 15 +/- 2% of the free L-Ser pool. The D- and L-Ser specific content and the D/L-Ser ratio obtained from temporal cortex of Parkinson and Alzheimer brains did not differ significantly from those of controls. However, at the levels observed here, and considering its specificity and affinity for the NMDA-associated glycine receptor, endogenous D-Ser is a plausible NMDA receptor glycine site agonist.

Pharmacokinetics and biochemical efficacy of pirmagrel, a thromboxane synthase inhibitor, in renal allograft recipients.

The effects of a 48-hour 0.5 mg/kg/hr infusion of the thromboxane synthase inhibitor pirmagrel were studied in 10 renal allograft recipients with cyclosporine nephrotoxicity. Plasma concentrations reached a mean steady-state plasma level of 1798 +/- 481 ng/ml. Biphasic, rapid elimination of pirmagrel was observed with a distribution half-life of 6.7 minutes and a terminal half-life of 73 minutes. Plasma clearance and the volume of distribution of the drug were 300 +/- 87 ml/hr/kg and 497 +/- 232 ml/kg, respectively. The pharmacodynamic effects of pirmagrel were marked by a mean 96% suppression of serum thromboxane B2 (TXB2), which coincided with a suppression of urinary excretion of TXB2, 2,3-dinor-TXB2, and 11-dehydro-TXB2 of 85% +/- 8%, 91% +/- 5%, and 89% +/- 9%, respectively. Urinary excretion of all thromboxane metabolites measured at the end of 1 week after termination of infusion was returned to the baseline. In conclusion, pirmagrel caused effective and sustained suppression of all thromboxane derived metabolites in plasma and urine during continuous infusion in kidney transplant patients receiving cyclosporine.

ACTH-induced increases in rabbit adrenal immunoreactive P-450(17 alpha) and P-450(21).

Rabbit adrenal 17 alpha-hydroxylase activity has previously been shown to increase dramatically following ACTH stimulation. The present study was designed to determine whether the increase in enzyme activity could be correlated with an increase in P-450(17 alpha) protein measured by immunoblotting using an anti-porcine P-450(17 alpha) antibody. It was found that the total and specific contents of rabbit adrenal immunoreactive P-450(17 alpha) were increased 6- to 8-fold and 4-fold, respectively, after ACTH stimulation. The results were similar whether the detection system was 125I-labeled protein A or an alkaline phosphatase-conjugated second antibody. Corresponding increases in 17 alpha-hydroxylase activity were also observed but were slightly less than the increases in immunoreactive P-450(17 alpha), suggesting that not all of the protein was enzymatically active. Comparatively, immunoreactive P-450(21) was increased only 1.3-fold. Antibodies to porcine P-450(17 alpha) and bovine P-450(21) reacted monospecifically with the homologous rabbit and guinea pig proteins as judged by the detection of single bands on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Inhibition studies showed that in an assay using 125 micrograms per ml of microsomal protein ACTH-stimulated rabbit adrenal 17 alpha-hydroxylase activity was inhibited 72% at a 100 mg per ml concentration of the anti-porcine P-450(17 alpha); however, 47% inhibition was observed at the same concentration of anti-bovine P-450(21). Pre-immune IgG had no effect. Molecular weight, Mr, determinations by SDS-PAGE showed both rabbit and guinea pig P-450(17 alpha) to be 52 kDa; rabbit P-450(21), 54 kDa; and guinea pig P-450(21), 49 kDa.(ABSTRACT TRUNCATED AT 250 WORDS)