domoic-acid and 4-methylglutamic-acid

Neurite outgrowth is a fundamental step in establishing proper neuronal connections in the developing central nervous system. Dynamic control of outgrowth has been attributed to changes in growth cone Ca2+ levels in response to extracellular cues. Here we have investigated a possible role for Ca2+ permeable kainate (KA) receptors in regulating neurite outgrowth of nociceptive-like dorsal root ganglion (DRG) neurons. To identify KA receptor subunits likely to be involved, we used quantitative RT-PCR on acutely dissociated DRG and dorsal horn neurons. DRG neurons expressed more GluK1, particularly the GluK1b spice variant, than dorsal horn neurons. Conversely, dorsal horn neurons expressed more GluK2, particularly GluK2a, than DRG neurons. Further, an RNA editing assay indicated that the majority of GluK1 and GluK2 mRNA transcripts in DRG were unedited. Imaging Ca2+ transients following application of a KA receptor agonist to DRG and dorsal horn co-cultures revealed increases in intracellular Ca2+ in the growth cones of DRG neurons. In the majority of cases, this increase in Ca2+ was partly or completely blocked by Joro spider toxin (JSTX), an antagonist for Ca2+-permeable AMPA and KA receptors. Treatment of DRG/dorsal horn co-cultures with KA for 18 hours suppressed neurite outgrowth while application of the rapidly desensitizing KA receptor agonist SYM 2081, the competitive AMPA/KA receptor antagonist, CNQX, and JSTX or philanthotoxin enhanced neurite outgrowth and prevented KA effects on neurite outgrowth. Thus, Ca2+ entry through KA receptors at the growth cone of DRG neurons may be an important regulator of neurite outgrowth.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Adenosine Deaminase; Analysis of Variance; Animals; Calcium; Cells, Cultured; Dose-Response Relationship, Drug; Embryo, Mammalian; Excitatory Amino Acid Antagonists; Ganglia, Spinal; GAP-43 Protein; Gene Expression Regulation; GluK2 Kainate Receptor; Glutamates; Growth Cones; Kainic Acid; Membrane Proteins; Neurites; Neuromuscular Depolarizing Agents; Rats; Rats, Sprague-Dawley; Receptors, Kainic Acid; RNA Editing; RNA, Messenger; Sensory Receptor Cells; Spider Venoms; Subcellular Fractions

Drosophila melanogaster larval neuromuscular junctions (NMJs) serve as a model for synaptic physiology. The molecular sequences of the postsynaptic glutamate receptors have been described; however, the pharmacological profile has not been fully elucidated. The postsynaptic molecular sequence suggests a novel glutamate receptor subtype. Kainate does not depolarize the muscle, but dampens evoked EPSP amplitudes. Quantal responses show a decreased amplitude and area under the voltage curve indicative of reduced postsynaptic receptor sensitivity to glutamate transmission. ATPA, a kainate receptor agonist, did not mimic kainate's action. The metabotropic glutamate receptor agonist t-ACPD had no effect. Domoic acid, a kainate/AMPA receptor agonist, blocks the postsynaptic receptors without depolarizing the muscle. However, SYM 2081, a kainate receptor agonist, did depolarize the muscle and reduce the EPSP amplitude at 1 mM but not at 0.1 mM. This supports the notion that these are generally a quisqualate subtype receptors with some oddities in the pharmacological profile. The results suggest a direct postsynaptic action of kainate due to partial antagonist action on the quisqualate receptors. There does not appear to be presynaptic auto-regulation via a kainate receptor subtype or a metabotropic auto-receptor. This study aids in furthering the pharmokinetic profiling and specificity of the receptor subtypes.

Topics: Afferent Pathways; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Benzodiazepines; Cycloleucine; Dose-Response Relationship, Drug; Drosophila; Electrophysiology; Excitatory Postsynaptic Potentials; Glutamates; Kainic Acid; Larva; Neuromuscular Junction; Receptors, AMPA; Receptors, Glutamate; Receptors, Kainic Acid; Receptors, Metabotropic Glutamate; Synaptic Transmission

We studied the modulation of gamma-aminobutyric acid (GABA) release by activation of kainate receptor in rat whole hippocampal synaptosomes. Kainate (10-300 microM) inhibited [3H]GABA release in a concentration-dependent manner with an EC50 of 25 microM. This effect of kainate (30 microM) was prevented by the ionotropic non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 microM) and by the selective kainate receptor antagonist 5-nitro-6,7,8,9-tetrahydrobenzo(g)indole-2,3-dione-3-oxime (NS-102, 10 microM), but not by the selective non-competitive AMPA receptor antagonist 1-(4-aminophenyl)-4-methyl-7,8-methylenedioxy-5 H-2,3-benzodiazepine (GYKI 52466, 100 microM). Other kainate receptor agonists, such as domoic acid (0.3-10 microM) and (2S,4R)-4-methylglutamic acid (MGA, 0.3-3 microM), also inhibited [3H]GABA release in a concentration-dependent manner with EC50 values of 4.0 microM and 0.90 microM, respectively, whereas alpha-amino-3-hydroxy-5-methyl-4-oxazolepropionate (AMPA, 10-100 microM) was devoid of effect. These inhibitory effects of both domoic acid (3 microM) and MGA (1 microM) were antagonized by CNQX (10 microM). These results indicate that GABA release can be modulated directly by presynaptic high-affinity kainate heteroreceptors.

Topics: Animals; Excitatory Amino Acid Agonists; GABA Antagonists; gamma-Aminobutyric Acid; Glutamates; Hippocampus; In Vitro Techniques; Kainic Acid; Male; Rats; Rats, Wistar; Receptors, Kainic Acid; Synaptic Transmission; Synaptosomes