strychnine and Neuroblastoma

The critical role of α1-glycine receptor (α1-GLYRs) in pathological conditions such as epilepsy is well known. In the present study, structure-activity relations for a series of phenylalanine derivatives carrying selected hydrogen bond acceptors were investigated on the functional properties of human α1-GLYR expressed in Xenopus oocytes. The results indicate that one particular substitution position appeared to be of special importance for control of ligand activity. Among tested ligands (1-8), the biphenyl derivative (2) provided the most promising antagonistic effect on α1-GLYRs, while its phenylbenzyl analogue (5) exhibited the highest potentiation effect. Moreover, ligand 5 with most promising potentiating effect showed in-vivo moderate protection when tested in strychnine (STR)-induced seizure model in male adult rats, whereas ligand 2 with highest antagonistic effect failed to provide appreciable anti(pro)convulsant effect. Furthermore, ligands 2 and 5 with the most promising effects on human α1-GLYRs were examined for their toxicity and potential neuroprotective effect against neurotoxin 6-hydroxydopamine (6-OHDA). The results show that ligands 2 and 5 possessed neither significant antiproliferative effects, nor necrotic and mitochondrial toxicity (up to concentration of 50μM). Moreover, ligand 2 showed weak neuroprotective effect at the 50μM against 100μM toxic dose of 6-OHDA. Our results indicate that modulatory effects of ligands 2 and 5 on human α1-GLYRs as well as on STR-induced convulsion can provide further insights for the design of therapeutic agents in treatment of epilepsy and other pathological conditions requiring enhanced activity of inhibitory glycine receptors.

Topics: Animals; Anticonvulsants; Convulsants; Disease Models, Animal; Dose-Response Relationship, Drug; Glycine; HEK293 Cells; Humans; Ligands; Male; Membrane Potentials; Microinjections; Neuroblastoma; Oocytes; Oxidopamine; Patch-Clamp Techniques; Phenylalanine; Rats; Rats, Wistar; Receptors, Glycine; Seizures; Strychnine; Transduction, Genetic; Xenopus laevis

Using fluo-3 calcium imaging, we demonstrate that glycine induces an increase in intracellular calcium concentration ([Ca2+]i) in cortical oligodendrocyte progenitor (OP) cells. This effect results from a calcium entry through voltage-gated calcium channels (VGCC), as it is observed only in OP cells expressing such channels, and it is abolished either by removal of calcium from the extracellular medium or by application of an L-type VGCC blocker. Glycine-triggered Ca2+ influx in OP cells actually results from an initial depolarization that is the consequence of the activation of both the ionotropic glycine receptor (GlyR) and Na+-dependent transporters, most probably the glycine transporters 1 (GLYT1) and/or 2 (GLYT2) which are colocalized in these cells. Through this GlyR- and transporter-mediated effect on OP intrcellular calcium concentration [Ca2+]i, glycine released by neurons may, as well as other neurotransmitters, serve as a signal between neurons and OP during development.

Topics: Amino Acid Transport Systems, Neutral; Animals; Bicuculline; Biological Transport; Calcium; Calcium Channel Blockers; Calcium Channels; Carrier Proteins; Cell Communication; GABA Antagonists; Glycine; Glycine Agents; Glycine Plasma Membrane Transport Proteins; Neuroblastoma; Neurons; Nifedipine; Oligodendroglia; Rats; Sodium; Stem Cells; Strychnine; Tumor Cells, Cultured

Topics: Anesthetics, Local; Animals; Cell Line; Glioma; Guanidine; Guanidines; Hybrid Cells; Ion Channels; Kinetics; Mice; Neuroblastoma; Rats; Strychnine; Veratridine; Veratrine

Both glial and neuronal cells maintained in primary culture were found to accumulate [3H]GABA by an efficient "high-affinity" uptake system (apparent Km = 9 muM, Vmax = 0.018 and 0.584 nmol/mg/min, respectively) which required sodium ions and was inhibited by 1 mM ouabain. Strychnine and parachloromercuriphenylsulfonate (pCS) (both at 1mM) also strongly inhibited uptake of [3H]GABA, but metabolic inhibitors (2,4-dinitrophenol, potassium cyanide, and malonate) were without effect. Only three structural analogs of GABA (nipecotate, beta-alanine, and 2,4-diaminobutyrate) inhibited uptake of [3H]GABA, while several other compounds with structural similarities to GABA (e.g. glycine, L-proline, and taurine) did not interact with the system. The kinetic studies indicated presence of a second uptake (Km = 92 muM, Vmax = 0.124 nmol/mg/min) in the primary cultures containing predominantly glioblasts. On the other hand, only one of the neuronal cell lines transformed by simian virus SV40 appeared to accumulate [3H]GABA against a concentration gradient. Apparent Km of this uptake was relatively high (819 muM), and it was only weakly inhibited by 1 mM ouabain and 1 mM pCS. The structural specificity also differed from that of the uptake observed in the primary cultures. Significantly, non of the nontransformed continuous cell lines of either tumoral (glioma, C6; neuroblastoma, M1; M1NN) or normal (NN;I6) origin actively accumulated [3H]GABA. It is suggested that for the neurochemical studies related to GABA and requiring homogeneous cell populations, the primary cultures offer a better experimental model than the continuous cell lines.

Topics: 4-Chloromercuribenzenesulfonate; Animals; Biological Transport; Cell Line; Embryo, Mammalian; gamma-Aminobutyric Acid; Glioma; Kinetics; Mice; Neuroblastoma; Neuroglia; Neurons; Ouabain; Rats; Strychnine

Topics: 4-Chloromercuribenzenesulfonate; Amino Acids; Aminooxyacetic Acid; Aspartic Acid; Atropine; Biological Transport, Active; Clone Cells; Culture Techniques; Cyanides; Dinitrophenols; Glutamates; Kinetics; Neuraminidase; Neuroblastoma; Neuroglia; Sodium; Strychnine; Temperature