No abstract

No abstract

The transport kinetics of the excitatory sulphur-containing amino acid (SAA) transmitter candidates, L-cysteine sulphinate (L-CSA), L-cysteate (L-CA), L-homocysteine sulphinate (L-HCSA), and L-homocysteate (L-HCA), together with their plasma membrane carrier specificity, was studied in cerebrocortical synaptosome fractions by a sensitive high performance liquid chromatographic assay. A high affinity uptake system could be demonstrated for L-CSA (Km = 57 +/- 6 microM; Vmax = 1.2 +/- 0.1 nmol/min/mg protein) and L-CA (Km = 23 +/- 3 microM; Vmax = 3.6 +/- 0.1 nmol/min/mg protein), whereas L-HCSA (Km = 502 +/- 152 microM; Vmax = 6.1 +/- 1.3 nmol/min/mg protein) and L-HCA (Km = 1550 +/- 169 microM; Vmax = 10.3 +/- 1.1 nmol/min/mg protein) exhibited much lower affinity as transport substrates. In all cases, only a single, saturable Na(+)-dependent component of uptake could be identified, co-existing with a non-saturable, Na(+)-independent influx component. Plasma membrane carrier specificity of the SAAs was established following comparison with other high-affinity neurotransmitter systems. High-affinity L-CSA and L-CA transport and low-affinity L-HCSA and L-HCA transport demonstrate strong positive correlations in inhibition profiles when compared against each other or individually against the high-affinity transport of L-[3H]glutamate, L-[3H]aspartate, or D-[3H]aspartate. Moreover, the transport systems for the excitatory SAAs exhibited a negative correlation when compared in inhibition profiles with the high affinity transport of both [3H] gamma-aminobutyric acid (GABA) and [3H]taurine.(ABSTRACT TRUNCATED AT 250 WORDS)

A dose-dependent, saturable, and calcium-dependent release of gamma-[3H]aminobutyrate [( 3H]GABA) from cortical neurones and D-[3H]aspartate from cerebellar granule cells following stimulation by a range of L-enantiomers of neuroactive acidic sulphur amino acids has been demonstrated. Moreover, the sulphur amino acid-evoked release of the transmitter amino acids was found to be sensitive to the presence of both selective N-methyl-D-aspartate and quisqualate/kainate receptor antagonists. Following the recent demonstration of an endogenous location for several of the acidic sulphur amino acids and their excitotoxic involvement in several neuropathological states and coupled with the knowledge that many important CNS connections are still undefined as far as their excitatory transmitter or transmitters are concerned, the present findings are of immediate importance in the continued search for endogenous excitatory amino acid agonists in addition to glutamate and aspartate.

A detailed kinetic study of the inhibitory effects of L- and D-enantiomers of cysteate, cysteine sulphinate, homocysteine sulphinate, homocysteate, and S-sulpho-cysteine on the neuronal, astroglial and synaptosomal high-affinity glutamate transport system was undertaken. D-[3H] Aspartate was used as the transport substrate. Kinetic characterisation of uptake in the absence of sulphur compounds confirmed the high-affinity nature of the transport systems, the Michaelis constant (Km) for D-aspartate uptake being 6 microM, 21 microM and 84 microM, respectively, in rat brain cortical synaptosomes and primary cultures of mouse cerebellar granule cells and cortical astrocytes. In those cases where significant effects could be demonstrated, the nature of the inhibition was competitive irrespective of the neuronal versus glial systems. The rank order of inhibition was essentially similar in synaptosomes, neurons and astrocytes. Potent inhibition (Ki approximately Km) of transport in each system was exhibited by L-cysteate, and L- and D-cysteine sulphinate whereas substantially weaker inhibitory effects (Ki greater than 10-1000 times the appropriate Km value) were exhibited by the remaining sulphur amino acids. In general, inhibition: (i) was markedly stereospecific in favor of the L-enantiomers (except for cysteine sulphinate) and (ii) was found to decrease with increasing chain length. Computer-assisted molecular modelling studies, in which volume contour maps of the sulphur compounds were superimposed on those of D-aspartate and L-glutamate, demonstrated an order of inhibitory potency which was, qualitatively, in agreement with that obtained quantitatively by in vitro kinetic studies.