Na(+)-Ca2+ exchange activity in its reverse mode was demonstrated in cultured rat astrocytes. Combination of ouabain (1 mM) and monensin (20 microM) caused a marked increase in 45Ca2+ uptake in astrocytes. 45Ca2+ uptake was also stimulated by lowering the external Na+ concentration. Ouabain plus monensin-stimulated 45Ca2+ uptake was blocked by 3,4-dichlorobenzamil (IC50, 16 microM), an inhibitor of Na(+)-Ca2+ exchanger, but not by nifedipine (0.1 microM). The stimulated-45Ca2+ uptake was observed even in K(+)-free medium, and external K+ at 5-10 mM caused a 2.2-fold increase in the uptake. Microspectrofluorimetry using the Ca(2+)-sensitive dye fura-2 showed that ouabain plus monensin increased intracellular Ca2+ concentration in single astrocytes. The Ca2+ signal was dependent on external Ca2+ (EC50, 1.4 mM), and blocked by 20 microM 3,4-dichlorobenzamil, but not by Ca2+ channel blockers (Cd2+, 20 microM; Ni2+, 100 microM). Antiserum of cardiac Na(+)-Ca2+ exchanger recognized 160 and 120-135 kDa proteins on SDS-polyacrylamide gel electrophoresis of astrocyte homogenate. Northern blot analysis revealed the presence of mRNA for the exchanger protein in astrocytes. These findings indicate that Na(+)-Ca2+ exchanger which is modulated by K+ is present in cultured rat astrocytes.

In some cells, Ca2+ depletion induces an increase in intracellular Ca2+ ([Ca2+]i) after reperfusion with Ca2+-containing solution, but the mechanism for the reperfusion injury is not fully elucidated. Using an antisense strategy we studied the role of the Na+-Ca2+ exchanger in reperfusion injury in cultured rat astrocytes. When astrocytes were perfused in Ca2+-free medium for 15-60 min, a persistent increase in [Ca2+]i was observed immediately after reperfusion with Ca2+-containing medium, and the number of surviving cells decreased 3-5 days later. The increase in [Ca2+]i was enhanced by low extracellular Na+ ([Na+]0) during reperfusion and blocked by the inhibitors of the Na+-Ca2+ exchanger amiloride and 3, 4-dichlorobenzamil, but not by the Ca2+ channel antagonists nifedipine, Ca2+ and Ni2+. Treatment of astrocytes with antisense, but not sense, oligodeoxynucleotide to the Na+-Ca2+ exchanger decreased Na+-Ca2+ exchanger protein level and exchange activity. The antisense oligomer attenuated reperfusion-induced increase in [Ca2+]i and cell toxicity. The Na+-Ca2+ exchange inhibitors 3, 4-dichlorobenzamil and ascorbic acid protected astrocytes from reperfusion injury partially, while the stimulators sodium nitroprusside and 8-bromo-cyclic GMP and low [Na+]0 exacerbated the injury. Pretreatment of astrocytes with ouabain and monensin caused similar delayed glial cell death. These findings suggest that Ca2+ entry via the Na+-Ca2+ exchanger plays an important role in reperfusion-induced delayed glial cell death.

Diclofenac was more effective than betamethasone in preventing angiographic CME and BAB disruption after small-incision cataract surgery. Thus, nonsteroidal antiinflammatory agents should be considered for routine treatment of eyes having cataract surgery.

1 5-Hydroxytryptamine (5-HT) plays a role in the regulation of noradrenergic neurones in the brain, but the precise mechanism of regulation of noradrenaline (NA) release by 5-HTIA receptors has not been defined. The present study describes the effect of a highly potent and selective 5-HTlA receptor agonist,release in the hypothalamus using microdialysis in the freely moving rat. 2 Subcutaneous injection of MKC-242 (0.5 mg kg-') increased extracellular levels of NA and its metabolite, 3-methoxy-4-hydroxyphenylglycol, in the hypothalamus and hippocampus.3 The 5-HTlA receptor agonists, 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) (0.2 mg kg-') and buspirone (3 mg kg-') mimicked the effect of MKC-242 in increasing NA release in the hypothalamus. 4 The effects of MKC-242 and 8-OH-DPAT in the hypothalamus were antagonized by pretreatment with WAY100135 (10 mg kg-'), a silent 5-HTlA receptor antagonist.5 Local administration of 8-OH-DPAT (10-100 /iM), citalopram (1 pM), a 5-HT reuptake inhibitor, and MDL72222 (10 pM), a 5-HT3 receptor antagonist, into the hypothalamus, had no effect on NA release. 6 Intracerebroventricular injection with 5,7-dihydroxytryptamine caused a marked reduction in brain 5-HT content, but the treatment affected neither basal NA levels nor the MKC-242-induced increase in NA release. 7 The effect of MKC-242 in increasing NA release was not attenuated by repeated treatment with the drug (0.5 mg kg-', once a day for 2 weeks). 8 The present results suggest that activation of postsynaptic 5-HTIA receptors increases NA release in the hypothalamus.

Although daily low-intensity pulsed ultrasound (LIPUS) can accelerate osteogenic differentiation of the rat clonal cell line ROS 17/2.8, the molecular mechanism that underlies this phenomenon is unclear. The purpose of this study was to determine which molecules exposed to daily LIPUS treatment stimulate osteogenic differentiation. The cells were cultured in the presence and absence (control) of LIPUS stimulation. LIPUS treatments consisted of 1.5-MHz ultrasound administered at an intensity of 30 mW/cm(2), 20 min daily for 7 days. The expression of bone morphogenetic proteins (BMPs) and their receptors involved in osteogenesis were measured using real-time PCR and/or Western blot analysis. Phosphorylation of the mothers against decapentaplegic 1 (Smad1) protein was determined by Western blotting. Daily LIPUS treatment significantly increased the expression of BMP-2, -4, and -7 and their receptors, and also phosphorylation of Smad1. Noggin markedly inhibited the daily LIPUS-induced phosphorylation of Smad1. Our findings demonstrate that the osteogenic activity of daily LIPUS may be mediated by BMPs in ROS 17/2.8 cells.