Amphiphysin I phosphorylation on residue threonine 260 in a pentylenetetrazole-induced seizure model

A method to evaluate kinase inhibitor action was reported [L. Morgan, S.J. Neame, H. Child, R. Chung, B. Shah, L. Barden, J.M. Staddon, T.R. Patel, Development of a pentylenetetrazole-induced seizure model to evaluate kinase inhibitor efficacy in the central nervous system, Neurosci. Lett. 395 (2006) 143–148]. In this, acute administration of the GABA antagonist pentylenetetrazole triggers seizures through glutamate-dependent pathways. Under such conditions, activation of the c-Jun N-terminal kinase (JNK) pathway was detected in hippocampal extracts. Phosphorylation of the upstream JNK kinase MKK4 was also revealed through use of a phospho-MKK4-specific antibody. Here, this antibody is shown to also react with a protein of ∼125 kDa which underwent increased phosphorylation in response to pentylenetetrazole treatment. The present study aimed to identify the ∼125 kDa protein as it may provide novel insight into signalling, neuronal activity and seizures. Using chromatographic methods and mass spectrometry, the protein was identified as amphiphysin I. This was confirmed by 2D gel analysis and immunoblot with amphiphysin I-specific antibodies. Although the phospho-MKK4 antibody was raised against an MKK4-specific peptide, partial sequence homology between this sequence and a region of amphiphysin was discerned. New antibodies raised against the phospho-threonine 260-amphiphysin-specific sequence detected increased phosphorylation in response to pentylenetetrazole treatment. This particular phosphorylation site does not seem to have been described before, possibly reflecting a novel regulatory aspect of amphiphysin biology. As amphiphysin is involved in the regulation of endocytosis, phosphorylation at this site may play a role in the regulated re-uptake of synaptic vesicles after neurotransmitter release.     

Cocaine and "pharmacological kindling" in the rat

The concept of "pharmacological kindling" has been used to explain the behavioral sensitization to cocaine produced by repeated administration of subconvulsive doses. This idea was tested by the repeated administration of cocaine to rats followed by electrical kindling of the olfactory bulb (a site at which cocaine has prominent electrophysiologic effects). No significant effect of cocaine on kindling was found. The relationship of this finding to studies using other drugs is discussed.     

The importance of free hydroxyl radicals to hypoxia preconditioning

Hypoxia preconditioning states that a sublethal hypoxia period will afford neuroprotection against a second harmful event. In our experiments, we carried out a procedure for the development of hypoxia preconditioning in adult male Wistar rats using hypoxic exposure (9% O(2); 91% N(2)) for 1 h. The protection against pentylenetetrazol (PTZ)-induced seizures was studied. For this, rats were tested by a single injection of PTZ (55 mg/kg i.p.) on days 1-21 after hypoxia exposure. The hypoxia exposure significantly prevented the development of acute PTZ convulsion at different times after hypoxia. The present study was designed to determine the effect of N-t-butyl-alpha-phenylnitrone (PBN), an electron-trapping agent and free radical scavenger, on hypoxia preconditioning against PTZ seizures 7 days after hypoxia exposure. PBN abolished the protective action of hypoxia exposure. The generation of free hydroxyl radicals in the brains of animals exposed to hypoxia was determined in a second experiment. For this purpose, the rats were i. p. pretreated with 30 mg/kg PBN and NaCl, respectively, 20 min before the start of hypoxia exposure. Forty-five minutes later the rats were i.p. injected with 300 mg/kg sodium salicylate and once again exposed to hypoxia for 15 min. Immediately after that the animals were decapitated and the free hydroxyl radicals and the salicylate content were estimated in the whole brain without cerebellum. Hypoxia preconditioned animals pretreated with NaCl showed a significantly higher extent of free hydroxyl radicals in the brain compared with PBN-injected preconditioned animals and with naive and sham exposed controls. The results pointed out that the generation of free reactive oxygen species under hypoxic conditions in the brain is involved in the development of the hypoxic preconditioning phenomenon.     

Effect of Pentylenetetrazole on Ethanol Intake, Ethanol Kinetics, and Social Behavior in Male Wistar Rats

Stress and anxiety are often implicated in excessive alcohol use. The nature of this interaction, however, is not understood. The aim of this study was to examine the effect of the anxiogenic agent, pentylenetetrazole (PTZ), on the acquisition and maintenance of ethanol drinking behavior in male Wistar rats. In rats maintained on a limited access procedure, with a choice between a 12% w/v ethanol (ETOH) solution and water available for 30 min each day, acute PTZ administration (1.5 to 15.0 mg/kg) did not modify ETOH intake. Chronic PTZ administration elicited a significant suppression in ETOH intake; however, this effect developed gradually over time. During the acquisition phase, chronic PTZ treatment also suppressed ETOH consumption. Chronic, but not acute, treatment with PTZ seemed to enhance water consumption. To assess whether the effect of PTZ on ETOH intake was due to either alterations in ETOH kinetics or behavior, blood ETOH levels and social interaction behaviour were examined. PTZ (15.0 mg/kg) produced a significant suppression in social interaction behavior, although tolerance developed to this effect on chronic PTZ administration. Both acute and chronic PTZ treatment (15 mg/kg) resulted in lower blood ETOH levels achieved after administration of 1.0 g/kg po of ETOH. Because the anxiogenic effect of PTZ was not maintained on repeated administration, yet the suppression of ETOH intake was only observed after chronic treatment, this suggests a dissociation between the processes regulating these behaviors.     

柴胡皂苷a对戊四氮诱导小鼠痫性发作的影响

目的:评价柴胡皂苷a(saikosaponin a,SSa)不同剂量对戊四氮(PTZ)诱导的小鼠痫性发作的影响。方法:采用PTZ诱导小鼠急性痫性发作来复制癫痫模型,通过记录小鼠痫性发作潜伏期和强直性惊厥率的变化,研究SSa抗小鼠PTZ致痫作用。结果:SSa高剂量组可以显著延长PTZ诱导的小鼠痫性发作(P<0.05),延长阵发性痉挛期向强直性惊厥发作的过渡时间(P<0.05),延长强直性惊厥发作的潜伏期(P<0.05),并且减少PTZ诱导小鼠强直性惊厥的发生率(P<0.05)。结论:SSa具有对抗小鼠PTZ致痫作用,对小鼠的强直性惊厥发生有较好的保护作用,并且存在量效关系的趋势。     

戊四氮慢性致痫大鼠海马区NF-kB、I-kBα动态表达及地塞米松的影响

目的:研究戊四氮慢性点燃大鼠即癫痫形成过程中海马区NF-Kb及其抑制蛋白I-kBα的动态变化及作用,并同时探讨地塞米松的抗痫作用机制.方法:将81只Wistar大鼠随机分为对照组(C),非用药组(P),用药组(D)即地塞米松干预组,除对照组外,均给予戊四氮(PIZ)35mg/kg·d腹腔注射.用免疫组织化学法检测不同时间点(4,7,11,15d)海马区NF-Kb、I-kBα的蛋白表达,同时行为学观察和脑电图确定痫性发作.结果:注射PIZ的各组大鼠海马区NF-Kb的表达随着注射时间逐渐增加,并于第15天达到高峰,且用药组相同时间点NF-Kb表达较非用药组减少(P<0.05); I-kBα蛋白量随着PTZ的注射逐渐减少(P<0.05),并于第15天降到最低,用药组相同时间I-kBα蛋白的减少较非用药组少(P<0.05);且地塞米松明显改善了大鼠点燃时的痫性发作程度,延长了发作潜伏期.结论:慢性癫痫的形成过程中海马区神经细胞中存在NF-Kb 的激活及其I-kBα的减少;NF-Kb介导了癫痫形成过程中的炎症反应过程,同时,地塞米松可能通过抑制NF-Kb活性发挥抗痫作用.     

托吡酯对戊四氮点燃慢性癫(癎)大鼠海马神经细胞黏附分子的影响

目的:探讨神经元活化在癫发生、发展中的作用及托吡酯对其的影响。方法:采用戊四氮制备慢性癫模型,利用托吡酯干扰,选取不同时间点,观察大鼠行为学变化及神经细胞黏附分子在海马回的表达(免疫组织化学染色)。结果:托吡酯组点燃率在27d明显低于模型组;模型组及托吡酯组神经细胞黏附分子表达增加,并随时间延长明显;而不同时间点该表达的增加程度,托吡酯组均低于模型组。结论:神经细胞黏附分子表达的增加,说明出现了神经元活化及脑可塑性变化,而托吡酯明显地抑制了该表达的增加。     

加减柴胡桂枝干姜汤对戊四唑诱导癫痫小鼠作用的实验研究

目的：研究加减柴胡桂枝干姜汤不同剂量对戊四唑诱导小鼠痫性发作的影响。方法：2月龄昆明种小白鼠连续灌胃给药7天后，腹腔注射戊四唑造成癫痫模型，造模后1h，观察药物对小鼠癫痫发作潜伏期、行为分级、发作次数等的影响。结果：加减柴胡桂枝干姜汤可以显著延长小鼠癫痫发作潜伏期、改善痫性发作程度、发作次数，与模型组相比，差异具有统计学意义（P〈0．05或P〈0．01），与西药组相比，无显著性差异（P〉0．05）。结论：加减柴胡桂枝干姜汤具有较好的抗小鼠戊四唑致痫作用，并存在一定的量效关系。     

Long-term theophylline treatment changes the effects of angiotensin II and adenosinergic agents on the seizure threshold

The effects of angiotensin II (ANG II), sarmesin, losartan, PD 123319, and adenosine A (1) receptor agonist N(6)-cyclopentyladenosine (CPA) administered i.c.v. in untreated and in theophylline-treated male mice (50 mg/kg i.p. twice daily for 14 days) were studied on the pentylenetetrazol (PTZ) seizure threshold. The threshold was increased after long-term theophylline treatment. ANG II, sarmesin, and CPA increased the threshold in theophylline-untreated mice, whereas it decreased the threshold in theophylline-treated animals. Losartan did not change the threshold in theophylline-untreated mice but decreased it in theophylline-treated animals. PD 123319 did not change the seizure threshold both in theophylline-untreated and -treated mice. Taken together, the data demonstrated that repeated exposure to theophylline selectively changes the effects of ANG II and adenosinergic agents on the PTZ seizure threshold. The results indicate that both angiotensin AT(1) and adenosine A(1) receptor subtypes could possess interactive mechanisms of adaptation to chronic theophylline treatment.