In vitro activation of soluble guanylyl cyclase and nitric oxide release: a comparison of NO donors and NO mimetics.

Nitric oxide (NO) performs a central role in biological systems, binding to the heme site of soluble guanylyl cyclase (sGC), leading to enzyme activation and elevation of intracellular levels of cGMP. Organic nitrates, in particular, nitroglycerin (GTN), are clinically important nitrovasodilators that function as NO-mimetics in biological systems. Comparison of sGC activation data with electrochemically measured rates of NO release for genuine NO donors, NONOates and nitrosothiols, yields an excellent correlation between the EC(50) for sGC activation and the rate constant for NO release, k(NO). However, activation of sGC by GTN and the nitrates has very different characteristics, including the requirement for specific added thiols, for example, cysteine. The reaction of GTN with cysteine in anaerobic solution yields NO slowly, and NO release, measured by chemiluminescence detection, is quenched by added metal ion chelator. The generation of NO under aerobic conditions is 100-fold slower than the anaerobic reaction. Furthermore, NO release from the reaction of GTN with cysteine in phosphate buffer is too slow to account for sGC activation by GTN/cysteine. The slow rate of the chemical reaction to release NO suggests that nitrates can activate sGC by an NO-independent mechanism. In contrast to the genuine NO donors, GTN behaves as a partial agonist with respect to sGC activation, but in the presence of the allosteric sGC activator, YC-1, GTN exhibits full agonist activity.

Modulation of AMPA receptors by a novel organic nitrate.

Positive modulators of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) channels reduce desensitization and alter their gating kinetics. We have discovered a novel compound nitric oxide-mimetic that similarly modulates the AMPA receptor by reducing desensitization. This, designated GT-005, belongs to the organic nitrate family that includes the nitrovasodilator nitroglycerine. In acutely isolated hippocampal neurons, GT-005 enhanced kainate (100 microM)-evoked currents with an EC50 of 1.7+/-0.2 mM and a 176+/-10% maximal increase in the steady-state current response. Similar results were found in cultured hippocampal neurons (EC50 of 1.3+/-0.2 mM and a maximal 83+/-14% increase in the steady-state current response). GT-005 reduced the desensitization of glutamate-evoked currents and slowed the onset of desensitization. This compound also increased the rate of recovery from the desensitized state. With respect to alteration of the excitatory synaptic transmission, GT-005 delayed the decay and increased the frequency of spontaneous miniature excitatory postsynaptic currents (mepsc) recorded in cultured hippocampal neurons.

Nitrate esters as nitric oxide donors: SS-nitrates.

[structure: see text]. The important biological secondary messenger NO can be generated from exogenous nitrovasodilators and NO donors. Nitrate esters are nitrovasodilators and NO mimetics, believed to be biotransformed to NO in vivo. On the basis of a mechanistic hypothesis, nitrates have been synthesized that release NO at significant rates in neutral aqueous solution in the presence only of added thiol. The novel masked beta-mercaptonitrates reported (SS-nitrates), provide information on possible sulfhydryl-dependent biotransformation mechanisms for nitrates in clinical use.