Altered activity–rest patterns in mice with a human autosomal-dominant nocturnal frontal lobe epilepsy mutation in the β2 nicotinic receptor

High-affinity nicotinic receptors containing β2 subunits (β2^*) are widely expressed in the brain, modulating many neuronal processes and contributing to neuropathologies such as Alzheimer's disease, Parkinson's disease and epilepsy. Mutations in both the α4 and β2 subunits are associated with a rare partial epilepsy, autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE). In this study, we introduced one such human missense mutation into the mouse genome to generate a knock-in strain carrying a valine-to-leucine mutation β2V287L. β2^(V287L) mice were viable and born at an expected Mendelian ratio. Surprisingly, mice did not show an overt seizure phenotype; however, homozygous mice did show significant alterations in their activity–rest patterns. This was manifest as an increase in activity during the light cycle suggestive of disturbances in the normal sleep patterns of mice; a parallel phenotype to that found in human ADNFLE patients. Consistent with the role of nicotinic receptors in reward pathways, we found that β2^(V287L) mice did not develop a normal proclivity to voluntary wheel running, a model for natural reward. Anxiety-related behaviors were also affected by the V287L mutation. Mutant mice spent more time in the open arms on the elevated plus maze suggesting that they had reduced levels of anxiety. Together, these findings emphasize several important roles of β2^* nicotinic receptors in complex biological processes including the activity–rest cycle, natural reward and anxiety

Effect of Adenosine A2A Receptor Antagonists and l-DOPA on Hydroxyl Radical, Glutamate and Dopamine in the Striatum of 6-OHDA-Treated Rats

A2A adenosine receptor antagonists have been proposed as a new therapy of PD. Since oxidative stress plays an important role in the pathogenesis of PD, we studied the effect of the selective A2A adenosine receptor antagonists 8-(-3-chlorostyryl)caffeine (CSC) and 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM 241385) on hydroxyl radical generation, and glutamate (GLU) and dopamine (DA) extracellular level using a microdialysis in the striatum of 6-OHDA-treated rats. CSC (1 mg/kg) and ZM 241385 (3 mg/kg) given repeatedly for 14 days decreased the production of hydroxyl radical and extracellular GLU level, both enhanced by prior 6-OHDA treatment in dialysates from the rat striatum. CSC and ZM 241385 did not affect DA and its metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanilic acid (HVA) extracellular levels in the striatum of 6-OHDA-treated rats. l-DOPA (6 mg/kg) given twice daily for two weeks in the presence of benserazide (3 mg/kg) decreased striatal hydroxyl radical and glutamate extracellular level in 6-OHDA-treated rats. At the same time, l-DOPA slightly but significantly increased the extracellular levels of DOPAC and HVA. A combined repeated administration of l-DOPA and CSC or ZM 241385 did not change the effect of l-DOPA on hydroxyl radical production and glutamate extracellular level in spite of an enhancement of extracellular DA level by CSC and elevation of extracellular level of DOPAC and HVA by ZM 241385. The data suggest that the 6-OHDA-induced damage of nigrostriatal DA-terminals is related to oxidative stress and excessive release of glutamate. Administration of l-DOPA in combination with CSC or ZM 241385, by restoring striatal DA-glutamate balance, suppressed 6-OHDA-induced overproduction of hydroxyl radical

Characterizing Ligand-Gated Ion Channel Receptors with Genetically Encoded Ca++ Sensors

We present a cell based system and experimental approach to characterize agonist and antagonist selectivity for ligand-gated ion channels (LGIC) by developing sensor cells stably expressing a Ca2+ permeable LGIC and a genetically encoded Förster (or fluorescence) resonance energy transfer (FRET)-based calcium sensor. In particular, we describe separate lines with human α7 and human α4β2 nicotinic acetylcholine receptors, mouse 5-HT3A serotonin receptors and a chimera of human α7/mouse 5-HT3A receptors. Complete concentration-response curves for agonists and Schild plots of antagonists were generated from these sensors and the results validate known pharmacology of the receptors tested. Concentration-response relations can be generated from either the initial rate or maximal amplitudes of FRET-signal. Although assaying at a medium throughput level, this pharmacological fluorescence detection technique employs a clonal line for stability and has versatility for screening laboratory generated congeners as agonists or antagonists on multiple subtypes of ligand-gated ion channels. The clonal sensor lines are also compatible with in vivo usage to measure indirectly receptor activation by endogenous neurotransmitters

The indirect basal ganglia pathway in dopamine D2 receptor-deficient mice

Recent pathophysiological models of basal ganglia function in Parkinson's disease predict that specific neurochemical changes in the indirect pathway would follow the lack of stimulation of D2 dopamine receptors. Post mortem studies of the basal ganglia in genetically modified mice lacking functional copies of the D2 dopamine receptor gene allowed us to test these predictions. When compared with their congenic N5 wild-type siblings, mice lacking D2 receptors show an increased expression of enkephalin messenger RNA in the striatum, and an increased activity and expression of cytochrome oxidase I in the subthalamic nucleus, as expected. In addition, D2 receptor-deficient mice display a reduced expression of glutamate decarboxylase-67 messenger RNA in the globus pallidus, as the basal ganglia model predicts. This reduction contrasts with the lack of change or increase in glutamate decarboxylase-67 messenger RNA expression found in animals depleted of dopamine after lesions of the mesostriatal dopaminergic system. Furthermore, D2 receptor-deficient mice show a significant decrease in substance P messenger RNA expression in the striatonigral neurons which form the direct pathway. Finally, glutamate decarboxylase-67 messenger RNA expression in the basal ganglia output nuclei was not affected by mutations in the D2 receptor gene, a fact that could probably be related to the absence of a parkinsonian locomotor phenotype in D2 receptor-deficient mice. In summary, these findings provide compelling evidence demonstrating that the lack of endogenous stimulation of D2 receptors is sufficient to produce subthalamic nucleus hyperactivity, as assessed by cytochrome oxidase I histochemistry and messenger RNA expression, and strongly suggest the existence of interactions between the basal ganglia direct and indirect pathways. (C) 2000 IBRO.Fil: Murer, Mario Gustavo. Inserm; Francia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Fisiología y Biofísica Bernardo Houssay. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Fisiología y Biofísica Bernardo Houssay; ArgentinaFil: Dziewczapolski, G.. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Farmacológicas. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Instituto de Investigaciones Farmacológicas; ArgentinaFil: Salin, P.. Centre National de la Recherche Scientifique; FranciaFil: Vila, M.. Inserm; FranciaFil: Tseng, Kuei y. Universidad de Buenos Aires. Facultad de Medicina; ArgentinaFil: Ruberg, M.. Inserm; FranciaFil: Rubinstein, Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Kelly, M. A.. University of Oregon; Estados UnidosFil: Grandy, D. K.. University of Oregon; Estados UnidosFil: Low, M. J.. University of Oregon; Estados UnidosFil: Hirsch, E.. Inserm; FranciaFil: Raisman Vozari, Rita. Inserm; FranciaFil: Gershanik, Oscar Samuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Ciencias Biológicas; Argentin