Impairment of neuropsychological behaviors in ganglioside GM3-knockout mice.

The ganglioside GM3 synthase (SAT-I), encoded by a single-copy gene, is a primary glycosyltransferase for the synthesis of complex gangliosides. Although its expression is tightly controlled during early embryo development and postnatal development and maturation in the brain, the physiological role of ganglioside GM3 in the regulation of neuronal functions has not been elucidated. In the present study, we examined motor activity, cognitive and emotional behaviors, and drug administration in juvenile GM3-knockout (GM3-KO) mice. GM3-KO male and female mice showed hyperactivity in the motor activity test, Y-maze test, and elevated plus maze test. In the Y-maze test, there was significantly less spontaneous alternation behavior in GM3-KO male mice than in wild-type mice. In the elevated plus maze test, the amount of time spent on the open arms by GM3-KO male mice was significantly higher than that of sex-matched wild-type mice. In contrast, there was no significant difference between GM3-KO and wild-type female mice in these tests. Thus, juvenile GM3-KO mice show gender-specific phenotypes resembling attention-deficit hyperactivity disorder (ADHD), namely hyperactivity, reduced attention, and increased impulsive behaviors. However, administration of methylphenidate hydrochloride (MPH) did not ameliorate hyperactivity in either male or female GM3-KO mice. Although these data demonstrate the involvement of ganglioside GM3 in ADHD and the ineffectiveness of MPH, the first-choice psychostimulant for ADHD medication, our studies indicate that juvenile GM3-KO mice are a useful tool for neuropsychological studies.

Enhanced CaMKII activity and spatial cognitive function in SAMP6 mice.

Senescence-accelerated mouse prone 6 (SAMP6) mice exhibit increased expression of NMDA receptor NR2B subunit (NR2B) and improved short-term memory compared with senescence-accelerated mouse resistance 1 (SAMR1) mice. The Thr286 phosphorylation of alpha calcium/calmodulin-dependent protein kinase II (CaMKII) has a crucial role in plasticity and learning among multiple downstream signaling pathways linked to the NMDA receptor. To examine the relationship between CaMKII activity and spatial learning in SAMP6, the authors employed western blot analysis and behavioral analyses (object location and delayed spatial win-shift eight-arm radial-maze tests). The levels of Thr286 and Ser831 phosphorylation of CaMKII and AMPA receptor subunit glutamate receptor 1 (CaMKII substrate), respectively, were increased in hippocampus of SAMP6 compared with SAMR1. SAMP6 showed faster hippocampal-dependent spatial memory formation than SAMR1 in both the object location and win-shift eight-arm radial-maze tests. Our results indicate that increased CaMKII activity influences the NR2B/CaMKII signal pathway and cognitive function in SAMP6.

Analysis of motor function and dopamine systems of SAMP6 mouse.

The motor function of senescence-accelerated mouse prone 6 (SAMP6) was evaluated with a battery of behavioral tests: locomotor activity test, traction test, wire hanging test, and rotating rod test. SAMP6 exhibited increased locomotor activity compared with senescence-accelerated mouse resistant 1 (SAMR1). There was no difference between SAMP6 and SAMR1 in the traction and wire hanging tests. In the rotating rod test, shorter retention times at each day in the accelerating version of the test were observed in SAMP6 compared with SAMR1, indicating a motor coordination deficit of SAMP6. To understand the mechanism involved, we focused on the dopamine system. Measurement of dopamine and its metabolites with HPLC revealed that the concentrations of dopamine in nucleus accumbens (NAcs) and cerebellum and of one or more dopamine metabolites in all tissues assayed were significantly higher in SAMP6 compared with SAMR1. Increases of dopamine transporter and dopamine receptor 1 (D1) in striatum, of dopamine receptor 3 (D3) in NAc, and of D1 and D3 in cerebellum in SAMP6 were observed. These results indicate that increased dopamine concentration in NAc and increased expression of D1 in striatum are possible cause(s) of the increased locomotor activity of SAMP6, and that increased D3 expression in cerebellum contributes to the motor coordination deficit of SAMP6.

Emotional behavior and expression patterns of tyrosine hydroxylase and tryptophan hydroxylase in senescence-accelerated mouse (SAM) P6 mice.

Senescence-accelerated prone mouse 6 (SAMP6) is a model for senile osteoporosis. It was recently reported that SAMP6 has a memory deficit in the water maze test. Because emotion and cognition are thought to interact, in the present study to examine emotional behavior in SAMP6, we employed a battery of tasks (open field, elevated plus maze, light-dark exploration, marble-burying behavior, tail suspension), using three age groups (1, 4, and 8 months of age) of SAMP6 mice and age-matched control SAMR1 (senescence-accelerated resistant mouse 1) mice. All three age groups of SAMP6 showed higher activity than SAMR1 in the open field test and reduced anxiety as measured in terms of time spent on the open arms in the elevated plus maze, time spent in the light box in the light-dark exploration, and time spent in marble-burying behavior in the marble-burying test. All three age groups of SAMP6 showed reduced immobility time compared with SAMR1 in the tail suspension test. Western blot analyses showed increased expression levels of tyrosine hydroxylase phosphorylated at serine-40 in striatum and nucleus accumbens and of tryptophan hydroxylase phosphorylated at serine-58 in brain stem of 1-month-old SAMP6. These results suggest that one possible reason for the alterations of motor activity and emotional behavior of SAMP6, at least after 1 month of age, is increased dopamine and serotonin levels.

Additive subthreshold dose effects of cannabinoid CB(1) receptor antagonist and selective serotonin reuptake inhibitor in antidepressant behavioral tests.

The main clinically used antidepressant drugs are selective monoamine reuptake inhibitors, including selective serotonin reuptake inhibitors (citalopram, sertraline), selective dopamine reuptake inhibitor (nomifensine) and selective noradrenaline reuptake inhibitor (reboxetine), but they have various side effects. Because cannabinoid CB(1) receptor antagonists (SR141716A, AM251) enhance monoamine release, they might be beneficial in the therapy of affective disorders. We hypothesized that the use of monoamine reuptake inhibitors in combination with cannabinoid CB(1) receptor antagonists would allow a lower dose of monoamine reuptake inhibitors to be used in the therapy of depression, thereby reducing or eliminating the side effects. To test this hypothesis, we examined the combination of SR141716A or AM251 with citalopram, sertraline, nomifensine or reboxetine at subthreshold doses to see whether these combinations would show an additive effect in the forced swimming test and the tail suspension test with mice. Subthreshold doses of cannabinoid CB(1) receptor antagonist and selective serotonin reuptake inhibitors, which separately had no effect on the immobility of mice in the tests, showed a clear effect when the drugs were administered at 40 and 30 min, respectively, before the tests, without any change of motor activity. Therefore, the use of subthreshold doses of these agents in combination might be useful to enhance mainly serotonergic neurotransmission, and to reduce or eliminate the side effects of citalopram and sertraline.

Age-related difference in nociceptive behavior between SAMP6 and SAMR1 strains.

Senescence accelerated prone mouse 6 (SAMP6) mice have been known to be a model for accelerated aging. Compared with the normal control senescence accelerated resistant mouse 1 (SAMR1) mice, although the SAMP6 mice have normal bone mass at 4 months, they exhibit a significantly lower bone mass at 8 months. It was recently reported that SAMP6 has memory deficit at 4 months of age, indicating that the change of nervous function might be already detected at 4 months of age. To assess whether SAMP6 mice exhibit an age-related abnormality of nociceptive transmission, we examined a battery of tests using the von Frey test for mechanically induced response, the hot plate test for thermally induced response, and the formalin paw test for chemically induced response. SAMP6 and SAMR1 showed similar response patterns in the von Frey test and the hot plate test. In the formalin paw test, 1-month-old SAMP6 and SAMR1 had similar responses, while 4-month-old SAMP6 exhibited attenuated phase 2 response, but normal phase 1 response. These findings indicate that onset of age-related phenotypes in SAMP6 differs in different tissues. SAMP6 could be useful to delineate the involvement of age-related nociceptive mechanisms.

Bovine ephemeral fever in Taiwan (2001-2002).

Bovine ephemeral fever (BEF), a vector-borne disease of cattle, is caused by the Ephemerovirus of the family Rhabdoviridae. In the past 40 years, Taiwan has had seven BEF epizootics, and we have previously reported the first five. This study summarizes the 2001 and 2002 epizootics; conducted case-control serologic studies on 10 herds involved in the 2001 epizootic; determined whether the recent BEF viruses have varied significantly; and discusses the relationship between epizootic patterns and possible variant BEF viruses. For mature cows that had received at least 2 doses of vaccine before the study, a negative correlation between the prevaccinated (the 3rd dose and after) serum neutralization antibody (SNA) titers and their postvaccinated peak rates was found. When prevaccinated SNA levels were at < or = 32, their postvaccinated SNA levels increased significantly faster (P<0.01) than for those at > or = 32. The glycoprotein gene of isolates from 1999, 2001, and 2002 had a 99.2-99.9% homology, without consistent amino acid variations in the neutralization sites. Phylogenetic analysis of Taiwanese isolates revealed 2 distinct clusters, the 1983-1989 and 1996-2002 isolates. Cross-neutralization tests confirmed the glycoprotein gene sequence analysis results. In conclusion, annual boosters at SNA levels > 32, at more than 2 doses, or at intervals shorter than 6 months are not advisable. The occurrence of frequent small epizootics implies the dominance of BEF virus over host immunity, but not a variant virus.

Sustained hypomyelination and high serum thyroid hormone in aged black tremor hamster.

Oligodendrocytes and myelin in the corpus callosum of black tremor and normal hamsters aged over 1.5 years were ultrastructurally examined to determine the myelination index (ratio of myelin thickness/diameter of axon), percentage of naked axons, and proportions of oligodendroglial subtypes (light, medium and dark). The mutant hamsters were remarkably hypomyelinated, with a low myelination index and a high proportion of naked axons, and high proportions of the dark subtypes. Serum concentrations of thyroid hormones (T3 and T4) in 6-week-old mutant hamsters were 2-fold (T3) to 3-fold (T4) higher than those of age-matched normal animals. However, in the aged animals (over 1.5 years old) only T4 levels of the mutant hamsters were higher in the mutant than normal hamsters. The black tremor hamsters were hypomyelinated throughout their life and high serum level of thyroid hormones might have played a role in the hypomyelination.

Rolling Nagoya mouse strain (PROD-rol/rol) with classic piebald mutation.

Ataxic rolling Nagoya (PROD-rol/rol) mice, which carry a mutation in the α1 subunit of the Cav2.1 channel (Cacna1a) gene, were discovered in 1969. They show white spots on agouti coat and have a mutation in the piebald spotting (s) locus. However, mutation analysis of the s locus encoding the endothelin receptor type B (Ednrb) gene in PROD-rol/rol mice had not been performed. Here, we examined the genomic and mRNA sequences of the Ednrb gene in PROD-rol/rol and wild-type rolling Nagoya (PROD-s/s) and studied the expression patterns of Ednrb and Cacna1a genes in these mice in comparison with C57BL/6J mice. Polymerase chain reaction analyses revealed two silent nucleotide substitutions in the coding region and insertion of a retroposon-like element in intron 1 of the Ednrb gene. Expression analyses demonstrated similar localizations and levels of Ednrb and Cacna1a expression in the colon between PROD-rol/rol and PROD-s/s mice, but the expression levels of both genes were diminished compared with C57BL/6J mice. Microsatellite genotyping showed that at least particular regions of chromosome 14 proximal to the Ednrb locus of the PROD strain were derived from Japanese fancy piebald mice. These results indicated that PROD-rol/rol mice have two mutant genes, Ednrb and Cacna1a. As no PROD strain had an intact Ednrb gene, using congenic rolling mice would better serve to examine rolling Nagoya-type Cav2.1 channel dysfunctions.

Age-dependent kainate sensitivity in heterozygous rolling Nagoya Cav2.1 channel mutant mice.

Cav2.1α1 is involved in glutamate release. The kainate-induced intensive firing of neurons via glutamate receptors causes seizure and neuronal damage, especially in the hippocampus. Cav2.1α1 mutation in homozygous rolling Nagoya (rol/rol) mice caused reduced Ca(2+) permeability compared to wild-type mice. The rol/rol mice exhibited ataxia approximately after 2 weeks of age. Although we have reported that heterozygous rolling Nagoya (rol/+) mice show age-dependent behavioral changes, sensitivity to kainate has not been examined. To examine the relationship between Cav2.1 function and neurological disease, we investigated how Cav2.1 is related to kainate-induced seizure and neuronal damage using 2- and 18-month-old rol/+ mice. The seizure scores of 18-month-old rol/+ mice that received 20mg/kg kainate intraperitoneally were significantly lower than those of wild-type mice. As a consequence of seizure, kainate induced delayed neuronal damage along with astrocytic growth in the hippocampus in wild-type mice, with a moderate effect observed in rol/+ mice. In the hippocampus of 18-month-old rol/+ mice, the levels of mutant-type Cav2.1α1 were increased compared to +/+ mice. The phosphorylation of p38, a mitogen-activated protein kinase (MAPK) activated by kainate, was not increased after kainate injection compared to +/+ mice. No difference was observed between 2-month-old rol/+ and wild-type mice intraperitoneally injected with 20mg/kg kainate in these analyses. These findings suggest that rol/+ mice experience age-related changes in sensitivity to kainate due to changes in the p38 MAPK signaling pathway via a mutant Cav2.1 channel. Hence, rol/+ mice may represent a novel model to delineate the association between Cav2.1 function, synaptic transmission, and the postsynaptic signaling cascade.

Improvement of spontaneous alternation behavior deficit by activation of α4ß2 nicotinic acetylcholine receptor signaling in the ganglioside GM3-deficient mice.

We have reported that in ganglioside GM3-deficient (GM3(-/-)) mice, spontaneous alternation behavior assessed by a Y-maze task was significantly lower, and total arm entries were significantly higher than in wild-type mice. The objective of the present study was to examine the role of nicotinic acetylcholine receptor (nAChR) signaling in impairment of spontaneous alternation behavior of GM3(-/-) mice. Nicotine treatment (0.3, 1.0 mg/kg, s.c.) dose dependently improved the spontaneous alternation deficit without affecting total arm entries in GM3(-/-) mice. The nicotine-induced (1.0 mg/kg, s.c.) improvement was significantly abolished by the nAChR antagonist mecamylamine (1.0 mg/kg, i.p.). The α4ß2 nAChR antagonist dihydro-ß-erythroidine (2.5, 10.0 mg/kg, i.p.) dose dependently counteracted the nicotine-induced improvement of spontaneous alternation in GM3(-/-) mice, whereas the α7 nAChR antagonist methyllycaconitine (2.5, 10.0 mg/kg, i.p.) did not. In addition, the α4ß2 nAChR agonist RJR-2403 (5.0, 10.0 mg/kg, s.c.) dose dependently and significantly improved the spontaneous alternation deficit, whereas the α7 nAChR agonist PNU120596 (0.3, 1.0, 3.0 mg/kg, i.p.) did not. These findings revealed that nicotine improved spontaneous alternation behavior of GM3(-/-) mice via the activation of α4ß2, but not α7, nAChR. Thus, the ganglioside GM3 might be responsible for α4ß2 nAChR signaling in the spontaneous alternation behavior.

Impaired motor function in senescence-accelerated mouse prone 1 (SAMP1).

Senescence-accelerated mouse prone (SAMP) strains of mice show early onset of senescence, whereas senescence-accelerated mouse resistant (SAMR) strains are resistant to early senescence and serve as controls. Although SAMP6 and SAMP8 are established models of central nervous system alterations, it is unclear whether SAMP1/Sku (SAMP1) is characterized by brain alterations and dysfunction related to behavioral functioning. In the present study, behavioral tests (i.e., locomotor activity, Y-maze, rotating rod, hind-limb extension, and traction), histochemistry, and Western blot analyses were employed to study this mouse model using 2- and 4-month-old SAMP1 and age-matched control SAMR1. Although 2-month-old SAMP1 and SAMR1 showed similar activity, 4-month-old SAMP1 exhibited less activity than age-matched SAMR1 in locomotor activity and Y-maze tests. In rotating rod test, 2- and 4-month-old SAMP1 showed motor-coordination dysfunction. An abnormal extension reflex in the hind-limb test was observed in 2- and 4-month-old SAMP1. There were no significant differences between SAMP1 and SAMR1 with respect to grip strength in the traction test or alternation behavior in the Y-maze test. Histochemistry and Western blot analyses exhibited that cerebellar Purkinje cells in 4-month-old SAMP1 mice persistently expressed tyrosine hydroxylase. These results suggest that SAMP1 is a useful model for examining mechanisms underlying motor dysfunction.

Ca2+ channel currents in dorsal root ganglion neurons of P/Q-type voltage-gated Ca2+ channel mutant mouse, rolling mouse Nagoya.

The role of the P/Q-type voltage-gated Ca(2+) channels (VGCCs) in release of neurotransmitters involved in nociception is not fully understood. Rolling mouse Nagoya (tg(rol)), a P/Q-type channel mutant mouse, expresses P/Q-type VGCC whose activation curve has a higher half activation potential and a smaller slope factor than the wild type channel. We previously reported that tg(rol) mice showed hypoalgesic responses to noxious stimuli. In this study, we examined the VGCC current in dorsal root ganglion (DRG) neurons by the whole-cell patch-clamp method. Both ω-agatoxin IVA (0.1 µM) and ω-conotoxin GVIA (1 µM) inhibited the VGCC current by about 40-50% in both the homozygous tg(rol) (tg(rol)/tg(rol)) and wild type (+/+) mice. The voltage-activation relationships of the total VGCC current and the ω-agatoxin IVA-sensitive component in the tg(rol)/tg(rol) mice shifted positively compared to the +/+ mice, whereas that sensitive to the ω-conotoxin GVIA was not different between the two genotypes. The time constant of activation of the VGCC current at -20 mV was longer in the tg(rol)/tg(rol) mice than in the +/+ mice. These changes in the properties of the VGCC in the tg(rol)/tg(rol) mouse may reduce the amount of the released neurotransmitters and account for the hypoalgesic responses.

Role of Ca(V)2.1-mediated NMDA receptor signaling in the nucleus accumbens in spatial short-term memory.

Heterozygous rolling Nagoya (rol/+) mice with a Ca(V)2.1α1 mutation show normal Y-maze behavior. Intra-accumbens injection of N-methyl-d-aspartate (NMDA; 0-2.0 µg/side) induced similar spontaneous alternations in wild-type and rol/+ mice; injections of NMDA receptor antagonist MK-801 (0.5 µg/side) or Ca(V)2.1 inhibitor levetiracetam (0.1 µg/side) did not affect controls but decreased spatial cognition in rol/+ mice, suggesting that Ca(V)2.1-mediated NMDA receptor signaling in the nucleus accumbens is involved in short-term spatial learning.

Emotional behavior in heterozygous rolling mouse Nagoya Ca v 2.1 channel mutant mice.

Although rolling mouse Nagoya, a Ca(v)2.1α(1) mutant, exhibits ataxia and elevated serotonin concentrations, heterozygous mice have not been examined in detail. Patients with heterozygous mutations in this orthologous gene exhibit neurological disorders. To examine the emotional behavior of heterozygous mice, we used behavioral tasks and examined Ca(v)2.1α(1) message levels, tryptophan hydroxylase expression patterns, and monoamine concentrations in 2- and 22-month-old mice. Reduced anxiety in the elevated plus maze, light-dark exploration, and marble-burying behavioral tests and reduced depression in the forced swimming and tail suspension tests were observed in 22-month-old heterozygous mice compared to aged-matched wild-type mice. The levels of mutant-type Ca(v)2.1α(1) message, phosphorylation of tryptophan hydroxylase, and serotonin increased in the brainstems of 22-month-old heterozygous mice. No difference was observed between 2-month-old heterozygous and wild-type mice in these analyses. These findings suggest that heterozygous mice show age-related emotional changes due to alterations in the serotonin system associated with mutant-type Ca(v)2.1α(1), and that heterozygous mice may represent a novel model to delineate the interaction between Ca(v)2.1 function and synaptic transmission.

Levetiracetam-mediated emotional behavior in heterozygous rolling Nagoya Ca(V)2.1 channel mutant mice.

Ca(V)2.1, which is highly expressed in the nervous system, plays an essential role in presynaptic neurotransmitter release. Although recent data suggest that the antiepileptic drug levetiracetam (LEV) inhibits presynaptic Ca(V)2.1 activity, the precise physiological role of Ca(V)2.1/LEV-regulated emotional performance has not been elucidated. We examined whether Ca(V)2.1/LEV mediates emotional behavior using a combined pharmacologic and genetic approach. Heterozygous rolling Nagoya (rol/+) mice carrying the Ca(V)2.1alpha(1) mutation demonstrated normal emotional behavior. Exposure to 75mg/kg LEV, which had no effect in wild-type controls, reduced anxiety in elevated plus maze and light-dark exploration tests and reduced depression in forced swimming and tail suspension behavioral tests in rol/+ mice. Similar behavioral patterns in motor activity were noted in wild-type and rol/+ mice injected with 0-150mg/kg LEV. The phosphorylation of tryptophan hydroxylase at serine-58 and serotonin concentration were increased in the brainstems of rol/+ mice injected with 75mg/kg LEV but not in those of wild-type controls. These results indicate that Ca(V)2.1/LEV mediates serotonin signaling leading to alterations in emotion. Our results also indicate that a combination of subthreshold pharmacologic and genetic approaches can be used to study functional signaling pathways in neuronal circuits.

Hallucinogenic 5-hydroxytryptamine 2A receptor agonist effects in senescence-accelerated mice.

Many neuropharmacological agents modulate the activity and conformation of heptahelical G protein-coupled receptors and activate ligand-specific signaling pathways. The hallucinogenic chemical 1-[2,5-dimethoxy-4-iodophenyl]-2-aminopropane (DOI), a serotonin receptor 2A (5-HT2AR) agonist, evokes extracellular signal-regulated kinase 1/2 (ERK1/2) signaling and head-twitch behavior. We previously reported that the senescence accelerated-prone mouse 6 (SAMP6) exhibited altered emotional behavior and increased levels of a serotonin-biosynthesizing enzyme compared to the senescence accelerated-resistant mouse 1 (SAMR1); however, the mechanism underlying the relationship between specific receptor signaling and behavioral phenotypes was unclear. In this study, we performed head-twitch tests and examined the total and phosphorylated levels of ERK1/2 and cAMP-responsive element-binding protein (CREB) in the bilateral somatosensory cortex to assess the differences between SAMP6 and SAMR1 using DOI. Although DOI dose-dependently increased the head-twitch response in both strains, the responses of SAMP6 given 0.3 and 1.0 mg/kg DOI were significantly greater than those of SAMR1 given DOI at the same doses. Although no dose-dependent increase in total ERK1/2 and total CREB expression was detected in response to DOI, the levels of phospho-ERK1/2 and -CREB increased in both strains. The phospho-ERK1/2 and -CREB levels in SAMP6 given 0.3 and 1.0 mg/kg DOI were significantly higher than those in SAMR1 given DOI at the same doses. These results indicate that SAMP6 increases DOI-dependent ERK1/2-CREB signaling leading to more head-twitch responses than SAMR1, and that SAMP6 could provide a useful model for examining the relationship between 5-HT2AR regulatory signaling and behavioral phenotypes.

Subthreshold pharmacological and genetic approaches to analyzing CaV2.1-mediated NMDA receptor signaling in short-term memory.

Ca(V)2.1 is highly expressed in the nervous system and plays an essential role in the presynaptic modulation of neurotransmitter release machinery. Recently, the antiepileptic drug levetiracetam was reported to inhibit presynaptic Ca(V)2.1 functions, reducing glutamate release in the hippocampus, although the precise physiological role of Ca(V)2.1-regulated synaptic functions in cognitive performance at the system level remains unknown. This study examined whether Ca(V)2.1 mediates hippocampus-dependent spatial short-term memory using the object location and Y-maze tests, and perirhinal cortex-dependent nonspatial short-term memory using the object recognition test, via a combined pharmacological and genetic approach. Heterozygous rolling Nagoya (rol/+) mice carrying the Ca(V)2.1alpha(1) mutation had normal spatial and nonspatial short-term memory. A 100mg/kg dose of levetiracetam, which is ineffective in wild-type controls, blocked spatial short-term memory in rol/+ mice. At 5mg/kg, the N-methyl-D-aspartate (NMDA) receptor blocker (+/-)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP), which is ineffective in wild-type controls, also blocked the spatial short-term memory in rol/+ mice. Furthermore, a combination of subthreshold doses of levetiracetam (25 mg/kg) and CPP (2.5mg/kg) triggered a spatial short-term memory deficit in rol/+ mice, but not in wild-type controls. Similar patterns of nonspatial short-term memory were observed in wild-type and rol/+ mice when injected with levetiracetam (0-300 mg/kg). These results indicate that Ca(V)2.1-mediated NMDA receptor signaling is critical in hippocampus-dependent spatial short-term memory and differs in various regions. The combination subthreshold pharmacological and genetic approach presented here is easily performed and can be used to study functional signaling pathways in neuronal circuits.

Impairment of spatial short-term memory following acute administration of the NMDA receptor antagonist in heterozygous rolling Nagoya mice carrying the Ca V 2.1 alpha1 mutation.

Heterozygous rolling Nagoya (rol/+) mice carrying Ca(V)2.1 alpha(1) mutation demonstrated normal behavior in Y maze test. Similar spontaneous alternation patterns were noted in wild-type and rol/+ mice injected with N-methyl-D-aspartate (NMDA; 0-50mg/kg, sc). Systemic injection of NMDA receptor blocker (MK-801; 0.05 mg/kg, ip) or intrahippocampal injection of MK-801 (0.5 microg/side), which had no effect in wild-type controls, decreased spatial cognition in rol/+ mice. These results indicate that Ca(V)2.1 alpha(1) mutation probably through decrease in Ca(2+) influx lowers the threshold for learning impairment. The combination subthreshold pharmacological and genetic approach is useful to study functional pathways in neuronal circuits.

Interaction between Ca(v)2.1alpha (1) and CaMKII in Ca (v)2.1alpha (1) mutant mice, Rolling Nagoya.

It has been reported earlier that interactions between Ca(v)2.1alpha(1) and calcium/calmodulin-dependent protein kinase II (CaMKII) in the presynaptic fraction and between the NMDA receptor subunit NR2B and CaMKII in the postsynaptic density (PSD) fraction are important for neuronal function. Ca(v)2.1alpha(1), CaMKII, and NR2B are predominantly expressed in the hippocampus. To examine the above interactions and CaMKII activity in the hippocampal presynapse and PSD of Rolling Nagoya mice carrying a mutation in Ca(v)2.1alpha(1) subunit, we performed immunoprecipitation and Western blot analyses. In the presynapse, the interaction between Ca(v)2.1alpha(1) and CaMKII and the phosphorylation of CaMKII (at Thr286) and its substrate Synapsin I (at Ser603) were decreased in mutant mice compared to wild-type mice. In the PSD, a similar pattern was observed for the interaction between NR2B and CaMKII and the phosphorylation of CaMKII (at Thr286) and its substrate AMPA receptor subunit glutamate receptor 1 (at Ser831) between mutant and wild-type mice. Our data indicate that disruption of the interaction between Ca(v)2.1alpha(1) and CaMKII may down-regulate presynaptic CaMKII activity and that Rolling Nagoya mice would be a useful model for examining presynaptic function.