Glutamic acid decarboxylase mRNA in the suprachiasmatic nucleus of rats housed in constant darkness

This study demonstrates that the levels of the mRNAs encoding the two isoforms of glutamic acid decarboxylase (GAD) (i.e., GAD65 and GAD67) do not differ over the circadian activity cycle in the suprachiasmatic nucleus (SCN) of rats housed in constant darkness. These data indicate that the rhythmic expression of GAD56 mRNA previously observed in animals housed in a light:dark cycle [K.L. Huhman, A.C. Hennessey, H.E. Albers, Rhythms of glutamic acid decarboxylase mRNA in the suprachiasmatic nucleus, J. Biol. Rhythms 11 (1996) 311-316.] is the result of the activity of retinal afferents.     

Valproate corrects the schizophrenia-like epigenetic behavioral modifications induced by methionine in mice.

BACKGROUND: Reelin and GAD(67) expression is downregulated in cortical interneurons of schizophrenia (SZ) patients. This downregulation is probably mediated by epigenetic hypermethylation of the respective promoters caused by the selective increase of DNA-methyltransferase 1 in GABAergic neurons. Mice receiving methionine (MET) provide an epigenetic model for neuropathologies related to SZ. We studied whether MET-induced epigenetic reelin promoter hypermethylation and the associated behavioral alterations can be reduced by valproate in doses that inhibit histone deacetylases (HDACs). METHODS: Mice treated with either methionine (MET) (5.2 mmol/kg/SC/twice daily) or valproate (1.5 mmol/kg/SC/twice daily) or MET+ valproate combination were tested for prepulse inhibition of startle (PPI) and social interaction (SI). S-adenosylmethionine, acetylated histone 3, reelin promoter methylation, and reelin mRNA were assayed in the frontal cortex. RESULTS: Valproate enhances acetylated histone 3 content, and prevents MET-induced reelin promoter hypermethylation, reelin mRNA downregulation, and PPI and SI deficits. Imidazenil, a positive allosteric modulator at GABA(A) receptors containing alpha(5) subunits but inactive at receptors including alpha(1) subunits, normalizes MET-induced behavioral changes. CONCLUSION: This MET-induced epigenetic mouse models the neurochemical and behavioral aspects of SZ that can be corrected by positively modulating the action of GABA at alpha(5)-containing GABA(A) receptors with imidazenil or by inhibiting HDACs with valproate, thus opening exciting new avenues for treatment of epigenetically modified chromatin in SZ morbidity.     

Decrease in GABA synthesis rate in rat cortex following GABA-transaminase inhibition correlates with the decrease in GAD67 protein

γ-Aminobutyric acid (GABA) synthesis in the brain is mediated by two major isoforms of glutamic acid decarboxylase, GAD₆₅ and GAD₆₇. The contribution of these isoforms to GABA synthesis flux (V_GAD) is not known quantitatively. In the present study we compared V_GAD in cortex of control and vigabatrin-treated rats under α-chloralose/70% nitrous oxide anesthesia, with total GAD activity and GAD isoform composition (GAD₆₅ and GAD₆₇) measured by enzymatic assay and quantitative immunoblotting. V_GAD was determined by re-analysis of ¹³C NMR data obtained ex vivo and in vivo during infusions of [1-¹³C]glucose using an extension of a model of glutamate–glutamine cycling that included a discrete GABAergic neuronal compartment with relevant interconnecting fluxes. V_GAD was significantly lower in vigabatrin-treated rats (0.030–0.05 μmol/min per g, P<0.003) compared to the non-treated control group (0.10–0.15 μmol/min per g). The 67–70% decrease in V_GAD was associated with a 13% decrease in total GAD activity (P=0.01) and a selective 44±15% decrease in GAD₆₇ protein (from 0.63±0.10 to 0.35±0.08 μg protein/mg tissue, P<0.05); GAD₆₅ protein was unchanged. The reduction in GAD₆₇ protein could account for a maximum of 65% of the decrease in V_GAD in vigabatrin-treated animals suggesting that inhibition of GAD₆₅ must have also occurred in these experiments, although product inhibition of GAD₆₇ by increased GABA could play a role. GAD₆₇ could account for 56–85% of cortical GABA synthesis flux under basal conditions and the entire flux after vigabatrin treatment.     

Repeated cocaine exposure during adolescence alters glutamic acid decarboxylase-65 (GAD65) immunoreactivity in hamster brain: correlation with offensive aggression

Male Syrian hamsters (Mesocricetus auratus) treated with low-dose (0.5 mg/kg/day) cocaine throughout adolescence (P27-P56) display highly escalated offensive aggression. The current study examined whether adolescent cocaine exposure influenced the immunohistochemical localization of glutamic acid decarboxylase-65 (GAD65), the rate-limiting enzyme in the synthesis of gamma-aminobutyric acid (GABA), a fast-acting neurotransmitter implicated in the modulation of aggression in various species and models of aggression. Hamsters were administered low doses of cocaine throughout adolescence, scored for offensive aggression using the resident-intruder paradigm, and then examined for changes in GAD65 immunoreactivity in areas of the brain implicated in aggression control. When compared with saline-treated control animals, aggressive cocaine-treated hamsters showed significant differences in the area covered by GAD65 puncta in several notable aggression regions, including the anterior hypothalamus, the medial and central amygdaloid nuclei, and the lateral septum. However, no differences in GAD65 puncta were found in other aggression areas, such as the bed nucleus of the stria terminalis, the ventrolateral hypothalamus, and the corticomedial amygdala. Together, these results suggest that altered GABA synthesis and function in specific aggression areas may be involved in adolescent cocaine-facilitated offensive aggression.     

A comparison of short-latency auditory-evoked potentials in two strains of mice: possible neurophysiological correlates of susceptibility to audiogenic seizures?

The short-latency auditory-evoked response was recorded from adult (over 8 wk) and young (16–22 day) mice of the audiogenic seizure-prone DBA/2J (D2) and seizure-resistant C57BL/6J (C6) strains. An auditory complex made up of eleven peaks and troughs was found within the first 20 ms after click stimuli. The latencies of the potentials tended to be shorter in the C6 mice and in the adults of both strains. The ratio of amplitude differences of late to early peaks (‘amplitude index’) was much larger(P<0.01) in D2 mice. The data are consistent with greater neuronal recruitment to acoustic stimuli in the higher-order auditory centers of D2 mice.     

Cortical spike wave discharges during audiogenic convulsions in rats

Audiogenic seizure-susceptible rats show high-amplitude spikes and waves in the electrocorticogram during the tonic phase of a maximal audiogenic convulsion. This observation establishes the validity of this seizure type as an animal model for human generalized epilepsy.     

Possible involvement of cytosolic phospholipase A(2) in cell death induced by 1-methyl-4-phenylpyridinium ion, a dopaminergic neurotoxin, in GH3 cells

Previously we reported that 1-methyl-4-phenylpyridinium ion (MPP(+)), a dopaminergic neurotoxin, induced apoptosis of GH3 cells established from rat anterior pituitary. In the present study, the role of MPP(+) along with that of other apoptotic factors such as Ca(2+) and H(2)O(2) in cell death was examined. Ionomycin induced DNA fragmentation and lactate dehydrogenase (LDH) leakage in GH3 cells. H(2)O(2) also induced LDH leakage. Co-addition of MPP(+), in conditions where MPP(+) had no effect by itself, enhanced ionomycin- and H(2)O(2)-induced cell death. Because the stimulation of phospholipase A(2) (PLA(2)) causing arachidonic acid (AA) release has been proposed to be involved in neuronal cell death, the effect of MPP(+) on AA release in GH3 cells was investigated. MPP(+) treatment for 8 h enhanced ionomycin- and H(2)O(2)-stimulated AA release mediated by activation of cytosolic PLA(2) in a concentration-dependent manner, although MPP(+) by itself had no effect on AA release. An inhibitor of cytosolic PLA(2) inhibited MPP(+)-induced cell death. These findings suggest a synergistic effect of MPP(+) on Ca(2+)- and H(2)O(2)-induced cell death, and the involvement of cytosolic PLA(2) activation in MPP(+)-induced cell death in GH3 cells. Pretreatment with a caspase inhibitor or EGF did not modify the ionomycin- or H(2)O(2)-induced AA release, or enhancement by MPP(+), but the pretreatment inhibited the cell death in the presence and absence of MPP(+). The involvement of caspase(s) on activation of PLA(2) by MPP(+) was excluded, and EGF inhibited MPP(+)-induced cell death downstream of the AA release.     

A Comparison of Cochlear Microphonics and N1 in Audiogenic-Seizure-Susceptible and Control Rats1

Abstract

Although numerous studies have shown that cochlear impainnent exists in audiogenic-seizure (AGS)-susceptible mice, there is only one report of cochlear potentials obtained from AGS-susceptible rats. To investigate the hypothesis that cochlear impairment also exists in AGS rats, cochlear microphonics (CM) and the primary afferent activity of the auditory division of the eighth cranial nerve (N₁) were studied in AGS rats.

AGS rats were obtained from the Veterans Administration Medical Center (Shreveport, La.) colony of Sprague Dawley derived animals, and control rats were obtained from Sprague Dawley, Inc. Two school bells ringing simultaneously were used to produce a sound of approximately 115 dB (AGS test stimulus). Exposure to the AGS test stimulus was once per week for three consecutive weeks. Chloramphenicol was used to treat the frequent otitis media found in the colony of AGS rats.

Two categories of AGS-susceptible and control rats were studied: (1) rats exposed to the AGS test stimulus and chloramphenicol regimen; and (2) rats not receiving these treatments. All rats were anesthetized with i.p. Dial-Urethane and prepared for cochlear round window recording. Cochlear microphonics were recorded in response to pure tones of 4, 8, 12, and 15 KHz, and N₁ was recorded in response to a click stimulus. A significant decrease in cochlear sensitivity was seen in both groups of AGS rats when compared to appropriate controls as reflected by a 25-35 dB shift in all CM and N₁ input-output functions. These results support the hypothesis that a functional cochlear impairment exists in the AGS rat.     

Divalent transition-metal ions (Cu and Zn) in the brains of epileptogenic and normal mice

The concentrations of Cu²⁺ and Zn²⁺ in 3 strains of mice were determined spectrophotometrically. The brain of the inborn audiogenic mouse (DBA/2J) contains higher levels of Zn²⁺ and Cu²⁺ than those found in the normal mouse (CBA/Ca or Parkes). Small differences in the metallic content in the whole brains of audiogenic and normal mice are accentuated in the hippocampus and the colliculus.     

Systemic Administration of N-Nitro-l-Arginine Methyl Ester and Indomethacin Reduces the Elevation of Brain PGE2Content and Prevents Seizures and Hippocampal Damage Evoked by LiCl and Tacrine in Rat

Administration of tacrine (5 mg/kg i.p.), an anticholinesterase agent, in rats pretreated (24 h beforehand) with lithium chloride (LiCl; 12 mEq/kg i.p.) enhances the expression of neuronal nitric oxide (NO) synthase (NOS), increases NO, and causes seizures and hippocampal damage. Here we report immunohistochemistry evidence showing that in rat LiCl and tacrine enhance the expression of cyclooxygenase type 2 (COX-2) enzyme protein in the dorsal hippocampus and elevate brain PGE₂content during the preconvulsive period. The latter effect, but not enhanced COX-2 expression, is inhibited by previous (30 min before tacrine) administration ofN^ω-nitro-l-arginine-methyl ester (L-NAME; 10 mg/kg i.p.), an inhibitor of NO synthesis, thus implicating NO in the mechanism of stimulation of COX activity leading to elevation of brain PGE₂content. Indomethacin (10 mg/kg given i.p. 30 min before tacrine), an inhibitor of COX activity, prevented brain PGE₂elevation and abolished the expression of seizures and hippocampal damage thus supporting a role for this metabolite of the arachidonic acid cascade in the mechanisms of LiCl and tacrine-evoked neurotoxicity in rat.     

Vitamin B(6) treatment of intractable seizures

Vitamin B₆ (VB₆)-related seizures include clinical seizures associated with VB₆ deficiency and dependency. Both types of seizures are suppressed by VB₆. We proposed VB₆-responsive seizures as the third category of VB₆-related seizures in 1977. Vitamin B₆-responsive seizures decrease or disappear in response to high-dose oral VB₆. Seizure onset in most of our cases occurred within the first year of life, although this varied between 3 months and 5 years. Etiologically, such cases were not only idiopathic or cryptogenic, but also symptomatic and associated with organic brain lesions. The tryptophan load test was usually negative. Vitamin VB₆-responsive seizures or epilepsy were usually West syndrome (WS), however may also include Lennox-Gastaut syndrome, grand mal or partial motor seizures. High-dose VB₆ treatment administered to 216 consecutive WS cases had an overall response rate of 13.9%, being high not only in cryptogenic cases (32%), but also in symptomatic WS (11.5%) associated with identifiable brain pathologies. Notably, responsive patients had excellent long-term seizure and mental outcomes without the need for conventional antiepileptic medication. A gradual increase in clinical response to VB₆ was noted with increasing the VB₆ dose from 30 to 50–100 mg/day, and a dramatic increase in clinical response with high-dose VB₆ (100–400 mg). Little clinical response was noted with administration of low dose VB₆ (10–30 mg/day). Thus, high-dose oral VB₆ treatment is recommended in all WS patients at time of initial treatment for a minimum of 10 days, considering the safety and rapid onset of efficacy, usually within 1 week, of this treatment.     

反复惊厥对P77PMC大鼠脑内BDNF蛋白表达的影响

采用免疫组织化学方法观察了反复听源性惊厥（ＡＣＳ）对遗传性癫痫易感大鼠Ｐ７７ＰＭＣ脑内脑源性神经营养因子（ＢＤＮＦ）蛋白表达的影响。结果发现,在未发生ＡＣＳ的Ｐ７７ＰＭＣ大鼠脑内,ＢＤＮＦ蛋白的基础表达可见于海马齿状回门区神经元、ＣＡ３区锥体细胞及大脑皮质神经元的胞浆内,在经过３０次ＡＣＳ发作后,鼠脑内齿状回门区、ＣＡ１－ＣＡ３区以及皮质区免疫标记阳性的的神经元数目增多,标记强度增强。除了胞体,在神经元突起上也可见明显的免疫标记。这表明在Ｐ７７ＰＭＣ这种遗传性癫痫易感大鼠模型中,非人为伤害因素造成的惊厥反复发作能导致脑内ＢＤＮＦ表达显著增高。     

建立胸部挤压全脑缺血大鼠慢性癫(癎)模型的操作细则

目的 建立一个简便、有效、稳定的脑缺血慢性癫(癎)模型,为探讨脑缺血与慢性癫(癎)的机理,及寻找新的癫(癎)药物治疗靶点提供基础和帮助.方法 采用胸部挤压方法造成大鼠脑缺血约6 min,立即予心肺复苏,24 h后予声音(闹铃110 dB)刺激诱导其癫(癎)发作,记录、评估癫(癎)发作情况,并行脑电图检查及分析.结果 采用胸部挤压构建大鼠缺血慢性癫(癎)模型的成功率为40％ ～ 60％,死亡率为30％,其发作类似于遗传性易感大鼠(GEPR)的音源性癫(癎)(AGS),其最典型的发作形式为狂奔,且脑电图特征符合癫(癎)的诊断.结论 胸部挤压全脑缺血大鼠慢性癫(癎)模型操作简便、制作成功率高、癫(癎)发作稳定可靠,且接近临床病理情况,可在研究脑缺血与慢性癫(癎)等研究中推广应用.     

Altered inhibitory input to Purkinje cells of dystrophin-deficient mice

We investigated evoked EPSPs and spontaneous IPSPs in cerebellar slices from dystrophin-deficient mdx mice. In the presence of the GABA(A) antagonist bicuculline the increase in EPSP amplitude was less in mdx Purkinje cells compared to control, and the amplitude of miniature IPSCs in mdx cells was also significantly less than in controls. This reduced inhibitory input is most likely due to the reported reduction in the size of GABA(A) channel clusters.