Adenosine modulates methylmercuric chloride (MeHgCl)-induced D-aspartate release from neonatal rat primary astrocyte cultures.

The effects of adenosine, and selective adenosine receptor agonists and antagonists on methylmercury (MeHg)-induced aspartate release were studied in neonatal rat primary astrocyte cultures. Whereas basal levels of D-[3H]aspartate release were unchanged upon treatment with adenosine or the selective A1 receptor agonists, N6-cyclopentyladenosine (CPA), cyclohexyladenosine (CHA), and R-phenylisopropyladenosine (R-PIA), all partially reversed the MeHg-induced release of D-aspartate. Treatment of astrocytes with the xanthine derivative, theophylline, an adenosine antagonist, reversed the inhibitory effect of adenosine on MeHg-induced D-[3H]aspartate release. Since the effect of MeHg on D-[3H]aspartate release is known to be associated with sulfhydryl (-SH) groups which are controlled by intracellular glutathione concentrations [GSH]i, we also evaluated the effects of adenosine, the A1 agonists CPA and CHP, and the adenosine antagonist, theophylline, on astrocytic [GSH]i. Attenuation of the stimulatory effect of MeHg on D-[3H]aspartate release by adenosine and its agonists occurred in the presence of reduced astrocytic [GSH]i, suggesting that other mechanisms must be invoked for this protective effect. Whilst the mechanism of MeHg-induced D-[3H]aspartate release is not known, the data suggest a role for adenosine in its regulation.

Astrocytes as potential modulators of mercuric chloride neurotoxicity.

1. MC has been shown to inhibit the uptake of L-glutamate and increase D-aspartate release from preloaded astrocytes in a dose-dependent fashion. 2. Two sulfhydryl (SH-)-protecting agents; reduced glutathione (GSH), a cell membrane-nonpenetrating compound, and the membrane permeable dithiothreitol (DTT), have been shown consistently to reverse the above effects. MC-induced D-aspartate release is completely inhibited by the addition of 1 mM DTT or GSH during the actual 5-min perfusion period with MC (5 microM); when added after MC treatment, DTT fully inhibits the MC-induced D-aspartate release, while GSH does not. 3. Neither DTT nor GSH, in the absence of MC, have any effect on the rate of astrocytic D-aspartate release. Other studies demonstrate that although MC treatment (5 microM) does not induce astrocytic swelling, its addition to astrocytes swollen by exposure to hypotonic medium leads to their failure to volume regulate. 4. Omission of calcium from the medium greatly potentiates the effect of MC on astrocytic D-aspartate release, an effect which can be reversed by cotreatment of astrocytes with the dihydropyridine Ca(2+)-channel antagonist nimodipine (10 microM), indicating that one possible route of MC entry into the cells is through voltage-gated L-type channels.

The role of sulfhydryl groups in D-aspartate and rubidium release from neonatal rat primary astrocyte cultures.

We have recently demonstrated that both methylmercury (MeHg) and mercuric chloride (MC) induce D-aspartate release from neonatal rat primary astrocyte cultures maintained in isotonic conditions. In the present study, we compare several other sulfhydryl-(-SH) selective alkylating reagents [methyl methanethiosulfonate (MMTS), N-ethylmaleimide (NEM), and iodoacetamide (IA)] in isotonic, as well as hypotonic conditions to discern the functional importance of -SH groups in [3H]D-aspartate and 86rubidium (86Rb) release from astrocytes. Treatment of astrocytes (5 min) in isotonic buffer with the hydrophobic reagent NEM (10 microM) caused a marked increase in 86Rb release but had no effect on [3H]D-aspartate release. Neither IA-, nor MMTS-treatment (both at 10 microM) induced increase in [3H]D-aspartate or 86Rb release in isotonic buffer. In hypotonic condition (-50 mM Na+), astrocytes were most sensitive to MC exposure (5 microM), exhibiting an increase in both [3H]D-aspartate and 86Rb efflux. The hydrophobic compounds MMTS and NEM, and the hydrophilic -SH modifying reagent, IA, attenuated the hypotonic-induced efflux of [3H]D-aspartate, in the absence of an effect on 86Rb release. These observations are consistent with a critical role for -SH groups both in basal (i.e. isotonic) and hypotonic-induced release of D-aspartate and Rb from astrocytes. Lack of uniformity of these effects may be attributed to site-specificity, related to the physicochemical properties of these -SH alkylating reagents.

The role of -SH groups in methylmercuric chloride-induced D-aspartate and rubidium release from rat primary astrocyte cultures.

Methylmercuric chloride (MeHgCl) was shown to increase D-aspartate and rubidium (Rb; a marker for potassium) release from preloaded astrocytes in a dose- and time-dependent fashion. Two sulfhydryl (-SH) protecting agents: a cell membrane non-penetrating compound, reduced glutathione (GSH), and the membrane permeable dithiothreitol (DTT), were found to inhibit the stimulatory action of MeHgCl on the efflux of radiolabeled D-aspartate as well as Rb. MeHgCl-induced D-aspartate and Rb release was completely inhibited by the addition of 1 mM DTT or GSH during the actual 5 min perfusion period with MeHgCl (10 microM). However, when added after MeHgCl treatment, this inhibition could not be fully sustained by GSH, while DTT fully inhibited the MeHgCl-induced release of D-aspartate. Neither DTT or GSH alone had any effect on the rate of astrocytic D-aspartate release. Accordingly, it is postulated that the stimulatory effect exerted by MeHgCl on astrocytic D-aspartate release is associated with vulnerable -SH groups located within, but not on the surface of the cell membrane. Omission of Na+ from the perfusion solution did not accelerate MeHgCl-induced D-aspartate release, suggesting that reversal of the D-aspartate carrier cannot be invoked to explain MeHgCl-induced D-aspartate release. Omission of Ca2+ from the perfusion solution increased the time-dependent MeHgCl-induced D-aspartate release.

Systolic blood pressure and long-term practice of the Transcendental Meditation and TM-Sidhi program: effects of TM on systolic blood pressure.

Systolic blood pressure was measured in 112 subjects practicing the Transcendental Meditation (TM) and TM-Sidhi programs. The subjects were between the ages of 35 and 64 years. A significant difference was found between the systolic blood pressures of subjects (matched for sex, race, and general educational background) practicing the TM and TM-Sidhi programs and norms for the general population. This difference was independent of diet and exercise patterns but related to length of time meditating. A significant difference was also found between short-term (under 5 years) and long-term (over 5 years) participants of the TM program, covarying for age. No previous reports exist concerning the long-term effects of the TM program on blood pressure. Despite methodological problems associated with cross sectional data, the findings suggest the beneficial effects of the long-term practice of the TM and TM-Sidhi programs on systolic blood pressure. Even if self-selection plays a role, the characteristics of an easily identifiable group already showing traits beneficial to the general population deserves further study.