Ion Channel Binding Sites Research Articles

Role of the NMDA receptor complex in DMCM-induced seizure in mice.

We investigated the role of the NMDA receptor complex in DMCM (methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate)-induced seizures in mice. The seizure threshold of DMCM was evaluated using an i.v. infusion technique. Pretreatment with the non-competitive NMDA receptor antagonist MK-801 ((+)-5-methyl-10,11-dihydro-5H-dibenzo(a,d) cycloheptan-5,10-imine maleate) or phencyclidine (PCP) significantly increased the seizure threshold for DMCM. Furthermore, the seizure threshold of DMCM was increased by intracerebroventricular (i.c.v.), but not intrathecal (i.t.), pretreatment with MK-801. Moreover, 7-chlorokynurenic acid, a glycine site antagonist, also increased the seizure threshold of DMCM, whereas ifenprodil, a non-competitive polyamine site antagonist, did not. These findings indicate that the ion-channel binding site and the glycine binding site on the NMDA receptor complex in the brain may be involved in the expression of seizures induced by DMCM.

Differential alterations of ion channel binding sites in temporal and occipital regions of the cerebral cortex in Alzheimer's disease

Three ion channel binding sites were examined by means of quantitative ligand binding autoradiography in temporal and occipital cortex from 9 patients with neuropathologically confirmed Alzheimer's disease (AD) and 7 matched control subjects. The following ligands were used: 125I-apamin to label a population of Ca 2+-sensitive K + channels; [ 3H]PN200-110 to label L-type voltage-sensitive Ca 2+ channels and [ 3H]glibenclamide to label ATP-sensitive K + channels. Ion channel binding sites were compared to: choline acetyltransferase (ChAT) activity and plaque densities measured in the same tissue. In the temporal cortex in AD 125I-apamin binding was increased compared to controls (e.g. superficial layers: control= 0.71 ± 0.07;AD= 1.02 ± 0.07,mean±S.E.M. pmol/g tissue). In contrast, in adjacent sections [ 3H]glibenclamide binding was reduced in AD compared to controls (e.g. superficial layers: control= 25.3 ± 1.7;AD= 17.9 ± 1.4pmol/g tissue). [ 3H]PN200-110 binding in temporal cortex was not altered in AD compared to controls. In the occipital cortex 125I-apamin binding was increased in AD while both [ 3H]glibenclamide and [ 3H]PN-200-110 binding sites in this cortical area were not different from controls. Plaque density (per mm 2) was higher in temporal (e.g. layers I–III, 43 ± 6) than in occipital cortex (layers I–III, 27 ± 4) in the AD patients while ChAT was reduced by 40% in temporal cortex and by 50% in occipital cortex compared to controls. The results suggests that the three ion channel binding sites are located on structural elements in the brain which are differentially affected by the pathophysiology of AD.

Effect of acute alcohol withdrawal on sensitivity to pro- and anticonvulsant treatments in WSP mice.

Through the genetic technique of selective breeding, a mouse line [Withdrawal Seizure Prone (WSP)] has been developed that expresses severe handling-induced convulsions (HIC) after cessation of chronic ethanol exposure. These mice also display rebound elevations in HIC after a single ethanol injection. In the current studies, we tested WSP mice in several paradigms. WSP mice were found to be marginally sensitive to the effects of acute doses of dizocilpine to reduce HIC. However, when tested during acute withdrawal from a single ethanol injection, WSP were more sensitive to this compound. Although N-methyl-D-aspartate significantly elevated HIC in naive WSP mice, it was more effective at low doses when given during acute withdrawal. Withdrawing mice were slightly more sensitive than naive mice to kainic acid. Pentylenetetrazole elevated HIC in naive and withdrawing mice; it was marginally more effective in naive mice. Diazepam inhibited HIC in both naive and withdrawing mice, and was slightly more effective during acute withdrawal. This pattern of results suggests that acute alcohol withdrawal is accompanied by altered sensitivity to convulsants and anticonvulsants. These changes include enhanced sensitivity in at least two excitatory amino acid-gated ion channel binding sites.

No Modifications of GABAA and Benzodiazepine Receptors Following Experimental Dysthyroidism in Rats

The effect of a treatment with L-triiodothyronine (T3) or propylthiouracil (PTU) on the characteristics of benzodiazepine and chloride ion channel binding sites in rat hippocampus and cerebral cortex was studied using a radiolabelled technique. In our experimental conditions, neither hyper- nor hypothyroidism modified number and affinity of [3H]flunitrazepam or [3H]butylbicycloorthobenzoate (TBOB) binding sites. These data indicate that neither benzodiazepine nor chloride ionophore sites of the GABA complex are modified in an experimental condition of dysthyroidism.

Postnatal changes in N-methyl-D-aspartate receptor binding and stimulation by glutamate and glycine of [3H]-MK-801 binding in human temporal cortex.

1. Homogenates of human infant and adult temporal cortex were used to measure [3H]-TCP and [3H]-MK-801 binding to the N-methyl-D-aspartate (NMDA)-coupled ion channel phencyclidine site. 2. Both [3H]-TCP and [3H]-MK-801 binding increased in infant cortex by > 100% between term and 26 weeks suggesting that the numbers of NMDA receptors increase during postnatal brain development. 3. [3H]-MK-801 binding was measured under non-equilibrium conditions in temporal cortex homogenates with the addition of 100 microM of L-glutamate plus a range of concentrations (0.05 microM-100 microM) of glycine. Glutamate and glycine increased [3H]-MK-801 binding by stimulating NMDA receptors and improving [3H]-MK-801 access to ion channel binding sites; maximum stimulation in adult and infant temporal cortex was achieved with 100 microM glutamate plus 5 microM glycine; a higher concentration of glycine (50 microM) reduced [3H]-MK-801 binding to below maximum. 4. The stimulation by 100 microM glutamate plus 5 microM glycine of [3H]-MK-801 binding in infant temporal cortex was affected by postnatal age. For example, although the stimulation of [3H]-MK-801 binding in 5-6 week infant cortex (236% of basal) was similar to adult cortex (230% of basal), in samples taken from infants aged 5-6 months glycine (plus glutamate) stimulation of [3H]-MK-801 binding (392% of basal) was substantially greater than that measured in adult temporal cortex. 5. The binding of [3H]-glycine to the glycine modulatory site associated with the NMDA receptor in infant cortex also increased with postnatal age by > 100% between term and 26 weeks. 6. It is concluded that NMDA receptors in infant cortex increase to levels greater than those in adult cortex during postnatal development. The results do not exclude the possibility that the transiently increased NMDA receptor-ion channel complex in infant cortex shows enhanced responses to agonists and modulators.

Multiple ion channel binding sites are differentially altered in Alzheimer's disease cortex: D. DEWAR, M. IKEDA, J. McCULLOCH. Wellcome Neuroscience Group, University of Glasgow, Glasgow, G61 1QH, U.K.

Multiple ion channel binding sites are differentially altered in Alzheimer's disease cortex: D. DEWAR, M. IKEDA, J. McCULLOCH. Wellcome Neuroscience Group, University of Glasgow, Glasgow, G61 1QH, U.K.

The quaking mouse: an epileptic mutant with alterations affecting the modulatory mechanisms of the NMDA receptor complex

The binding of [ 3H]glutamate and of [ 3H]1-(1-(2-thienyl)cyclohexyl)piperidine ([ 3H]TCP) has been examined in the genetically epileptic mutant mouse, quaking. The density of [ 3H]glutamate binding sites did not differ between the quaking mice and their controls of the same strain. In the absence of exogenous glutamate or glycine, the density of [ 3H]TCP binding sites was also similar in the two strains. In both the mutants and their controls, exogenously added glutamate, glycine and glutamate plus glycine dose-dependently increased the binding of [ 3H]TCP. In the 3 conditions, the modulation of [ 3H]TCP binding was significantly more efficient in the quaking mice than in the controls. Furthermore, in the presence of glutamate (10 −5 M), the increase of the affinity of the ligand for the ion channel binding site was higher in the mutants than in the controls. These results suggest that the modulatory mechanisms of the N- methyl- d-aspartate/ionophore receptor complex might be altered in these mutants. These alterations might be related to the previously observed anticonvulsant properties of NMDA receptor antagonists in the quaking mouse model of inherited epilepsy.

Role of hydrogen bonding in ligand interaction with the N-methyl-D-aspartate receptor ion channel

Displacement of [3H]MK-801 (dizocilpine, 1) binding to rat brain membranes has been used to evaluate the affinities of novel dibenzocycloalkenimines related to 1 for the ion channel binding site (also known as the phencyclidine or PCP receptor) on the N-methyl-D-aspartate (NMDA) subtype of excitory amino acid receptor. In common with many other agents having actions in the central nervous system, these compounds contain a hydrophobic aromatic moiety and a basic nitrogen atom. The conformational rigidity of these ligands provides a unique opportunity to evaluate the importance of specific geometrical properties that influence active-site recognition, in particular the role of the nitrogen atom in hydrogen-bonding interactions. The relative affinities (IC50s) of hydrocarbon-substituted analogues of 1 and ring homologated cyclooctenimines illustrate the importance of size-limited hydrophobic binding of both aryl rings and of the quaternary C-5 methyl group. Analysis of the binding of a series of the 10 available structurally rigid dibenzoazabicyclo[x.y.z]alkanes, by using molecular modeling techniques, uncovered a highly significant correlation between affinity and a proposed ligand-active site hydrogen bonding vector (r = 0.950, p less than 0.001). These results are used to generate a pharmacophore of the MK-801 recognition site/PCP receptor, which accounts for the binding of all of the known ligands.

Acetylcholine operated ion channel and α-Bungarotoxin binding site in a human neuroblastoma cell line reside on different molecules

In neurons, α-bungarotoxin is often associated with nicotinic receptor but does not always block the acetylcholine operated channel. In a human neuroblastoma cell line, IMR 32, we have demonstrated a large number of α-Bungarotoxin binding sites (2640 per cell in non differentiated cells and 4660 per cell in differentiated cells) in presence of 0 to 4 Acetylcholine activated-channels per cell. This neuronal cell line promises to be an useful model for the study of structure and function of the α-Bungarotoxin binding site not related to the nicotinic receptor.

Interactions of chlordecone (kepone) and mirex with the nicotinic acetylcholine receptor—ion channel complex

Interactions of chlordecone (kepone) and mirex with nicotinic acetylcholine (ACh) receptor complexes from Torpedo californica electric organs were studied using biochemical probes for ACh and ion-channel binding sites. Neither compound inhibited the binding of [ 125I]α-bungarotoxin (BGT) to the receptor; chlordecone, however, enhanced carbamylcholine affinity for the receptor 5-fold. Chlordecone, but not mirex, also inhibited the binding of [ 3H]perhydrohistrionicotoxin and [ 3H]phencyclidine to sites associated with the receptor-gated ion channel. Ion-channel inhibition by chlordecone was enhanced in the presence of carbamylcholine. These results indicate that chlordecone, but not mirex, interacts with the ion-translocation mechanism associated with nicotinic ACh receptors, where it may sterically block ion flux as well as stabilize a desensitized conformation of the receptor complex.