辣椒素受体参与骶髓后联合核神经元突触传递

目的研究辣椒素受体对大鼠骶髓后联合核(SDCN)神经元突触传递的影响。方法在脊髓骶段横切薄片上,利用全细胞膜片钳法记录骶髓后联合核神经元谷氨酸能兴奋性突触后电流(EPSCs)和γ-氨基丁酸(GABA)能抑制性突触后电流(IPSCs),比较激动辣椒素受体后上述突触电流的变化;观察激动辣椒素受体对SDCN神经元动作电位发放的影响。结果辣椒素受体被其特异性激动剂辣椒素(1μmol.L-1)激动后,自发EPSCs(sEPSCs)的频率和振幅均有明显增加(P<0.05,n=17)。在河豚毒素(0.5μmol.L-1)存在的条件下,辣椒素明显增加微小EPSCs(mEPSCs)的频率(P<0.01,n=13),但对mEPSCs的振幅无影响(P>0.05,n=13),提示辣椒素的作用在突触前。辣椒素也明显增加动作电位发放(P<0.05,n=19)。上述作用均可被辣椒素受体特异性拮抗剂capsazepine(10μmol.L-1)阻断。辣椒素也增加GABA能的自发IPSCs(sIPSCs)的频率(P<0.05,n=20),但对其不依赖动作电位的微小IPSCs(mIPSCs)的频率或振幅均无作用(P>0.05,n=9)。结论在SDCN,辣椒素受体主要表达于兴奋性突触终末;激动辣椒素受体影响兴奋性和抑制性突触活动,并可能参与痛觉信息在脊髓水平的传递和调制。     

SKF97541抑制大鼠骶髓后联合核神经元

目的研究GABAB受体特异性激动剂SKF97541对骶髓后联合核(SDCN)神经元的作用。方法在大鼠骶段脊髓横切薄片上,利用全细胞膜片钳法记录骶髓后联合核神经元。电流钳记录模式下,观察SKF97541对神经元膜电位和动作电位发放的影响。电压钳模式下,观察谷氨酸能兴奋性突触后电流(EPSCs)对SKF97541处理的变化。结果SKF97541(0.5μmol.L-1)通过作用于GABAB受体,减少SDCN神经元动作电位发放,同时促进细胞膜超极化。SKF97541在电压钳模式下,减少谷氨酸介导的微小EPSCs的频率,但对振幅无影响,提示SKF97541通过作用于突触前GABAB受体抑制谷氨酸释放。突触前刺激引起的突触后电位,也被SKF97541抑制。结论在骶髓后联合核,SKF97541通过作用于突触后GABAB受体,直接抑制神经元的兴奋性和动作电位发放;并通过突触前GABAB受体,抑制谷氨酸的释放。以上结果提示SKF97541的抑制作用可能抑制骶髓后联合核神经元对伤害性信息的传递。     

γ-氨基丁酸与甘氨酸受体在脊髓背角的分布及相互作用

外周伤害性刺激经脊髓、脑干和丘脑的传递和调制,最后在大脑皮层形成痛觉。脊髓在调制伤害性信息传递方面起着重要作用,脊髓背角是中枢神经系统痛觉信息整合加工的重要部位,由初级感觉传入末梢、脊髓中间神经元、脊髓投射神经元和脊髓内的下行纤维组成,构成复杂的神经网络,是感觉信息传人的门户和整合的初级中枢。     

丙泊酚对大鼠海马突触体释放谷氨酸和γ-氨基丁酸的影响

目的观察丙泊酚对大鼠海马突触体谷氨酸和γ氨基丁酸(GABA)Ca2+依赖性释放和非Ca2+依赖性释放的影响。方法制备突触体后用人工脑脊液(aCSF)孵育,分为7组,应用丙泊酚(propofol)的浓度分别为3(Pro3组)、10(Pro10组)、30(Pro30组)、100(Pro100组)和300μmol·L-1(Pro300组),脂肪乳剂对照组加入脂肪乳(Intralipid组),空白对照组(Control组)不加入任何药物。在观察Ca2+依赖性释放时,加入二氢海人藻酸和哌啶酸;在观察非Ca2+依赖性释放的时候,去除aCSF中的Ca2+。在37℃的条件下,应用20μmol·L-1藜芦定或30mmol·L-1KCl诱发递质释放。应用反相高效液相色谱法测定aCSF中谷氨酸和GABA的浓度。结果①Ca2+依赖性递质释放:应用藜芦定时,Pro30、Pro100和Pro300组的谷氨酸和GABA释放量低于Intralipid组(P<0.01或P<0.05);应用KCl时,各组的谷氨酸和GABA释放量无差异(P>0.05)。②非Ca2+依赖性释放:各组应用藜芦定和KCl诱发的谷氨酸和GABA释放量无差异(P>0.05)。结论丙泊酚可以抑制Ca2+依赖性谷氨酸和GABA的释放,而对非Ca2+依赖性谷氨酸和GABA释放无影响。     

γ-氨基丁酸受体系统在脑缺血诱发神经元再生过程中的调节作用

神经元再生已受到越来越多学科的重视。脑缺血性损伤可以诱发大脑生发中心神经干细胞的增殖、迁移并最终分化成神经元细胞。研究发现,脑缺血性损伤可以引起γ氨基丁酸(GABA)受体表达和功能的下调。GABA及其受体在神经元的发生和发育过程中具有营养、形态发生素等作用,同时还可以易化神经细胞形成。因此,GABA受体的变化可能与脑缺血性损伤诱发神经元再生密切有关。本文就GABA受体在脑缺血诱发神经元再生过程中调节作用的研究进展进行综述。     

丙泊酚对丘脑室旁核谷氨酸能神经元活性的影响

本研究利用全细胞膜片钳技术探索丙泊酚对丘脑室旁核(paraventricular thalamus, PVT)谷氨酸能神经元活性的影响及作用机制。在8周龄C57BL/6J小鼠急性脑片上,用单细胞逆转录PCR技术鉴定PVT神经元类型。记录丙泊酚给药前、后和洗脱后PVT神经元的放电频率(firing frequencies before, during, and after, FB, FD and FW)及给药前、后的膜电位(membrane potential before and during, MPB and MPD)。探索木防己苦毒素(picrotoxin, PTX)阻断γ-氨基丁酸A型(gamma-aminobutyric acid type A, GABAA)受体后对丙泊酚作用的影响,以及丙泊酚对PVT神经元上自发和微小抑制性突触后电流(spontaneous and miniature inhibitory postsynaptic currents, sIPSCs and mIPSCs)的影响。动物实验已获得复旦大学上海医学院动物实验伦理委员会批准。结果显示,在脂肪乳组和2...     

HPLC测定大鼠海马中谷氨酸、γ-氨基丁酸的含量

目的建立了邻苯二甲醛（OPA）柱前衍生高效液相荧光色谱法检测大鼠海马中谷氨酸（GLU）、γ-氨基丁酸（GABA）含量的方法。方法采用AlltimaC18色谱柱（4.6mm×250mm,5μm）,流动相为50mmol·L^-1乙酸钠（pH6.8）、甲醇、四氢呋喃梯度洗脱,流速1.0mL·min^-1。激发波长（λEx）338nm,发射波长（λEm）425nm。结果GLU、GABA线性范围分别为1.03～20.6μg·mL^-1（r=0.9992）和1.13～22.6μg·mL^-1（r=0.9997）;当信噪比为3时,GLU和GABA的检出限分别为5.27×10-4μg·mL^-1和7.35×10-4μg·mL^-1;测得精密度RSD分别为3.4%和3.5%;加样回收率分别为90%～96%,85%～91%。结论本方法简便、准确、灵敏,特异性强,重复性好,适用于海马中GLU和GABA的含量测定。     

HPLC测定大鼠海马中谷氨酸、γ-氨基丁酸的含量

目的 建立了邻苯二甲醛(OPA)柱前衍生高效液相荧光色谱法检测大鼠海马中谷氨酸(GLU)、γ-氨基丁酸(GABA)含量的方法。方法 采用Alltima C₁₈色谱柱(4.6 mm×250 mm,5 μm),流动相为50 mmol·L^-1乙酸钠(pH 6.8)、甲醇、四氢呋喃梯度洗脱,流速1.0 mL·min^-1。激发波长(λ_Ex)338 nm,发射波长(λ_Em)425 nm。结果 GLU、GABA线性范围分别为1.03~20.6 μg·mL^-1(r=0.999 2)和1.13~22.6 μg·mL^-1(r=0.999 7);当信噪比为3时,GLU和GABA的检出限分别为5.27×10^-4μg·mL^-1和7.35×10^-4 μg·mL^-1;测得精密度RSD分别为3.4%和3.5%;加样回收率分别为90%~96%,85%~91%。结论 本方法简便、准确、灵敏,特异性强,重复性好,适用于海马中GLU和GABA的含量测定。     

锌离子对大鼠骶髓后连合核神经元甘氨酸激活的全细胞电流的调控

目的研究锌离子对急性分离的大鼠骶髓后连合核神经元甘氨酸激活全细胞电流的调控。方法采用制霉菌素穿孔膜片钳方法。结果锌离子（10～1000μmol·L-1）呈浓度依赖性地抑制甘氨酸激活的全细胞电流。结论在大鼠骶髓后连合核，锌离子对甘氨酸激活的电流呈单相调控作用，即高浓度（≥10μmol·L-1）的锌离子抑制甘氨酸的反应，而低浓度（＜10μmol·L-1）的锌离子则无此作用。     

血浆谷氨酸与γ-氨基丁酸水平对麻醉后幼儿早期认知功能障碍的相关性研究

目的 观察丙泊酚或依托咪酯麻醉对幼儿腹腔镜下疝修补术后早期认知功能及血浆谷氨酸（Glu）与γ-氨基丁酸（GABA）含量的改变,探讨血浆谷氨酸与γ-氨基丁酸水平变化与丙泊酚或依托咪酯麻醉后幼儿早期认知功能障碍的相关性。方法 100例1³岁拟行择期腹腔镜下疝修补术患儿,随机分为丙泊酚麻醉组（A组）、依托咪酯麻醉组（B组）各50例;50例同年龄段健康幼儿作对照组（C组）。A、B组麻醉诱导分别给予丙泊酚1³ mg·kg^-1、依托咪酯0.1⁰.4 mg·kg^-1。比较A、B组内及组间术后认知功能障碍组和术后非认知功能障碍组患儿血浆谷氨酸、γ-氨基丁酸含量的变化。结果 A、B组术后3 d发生术后认知功能障碍的例数分别为10例（20.0%）、9例（18.0%,P>0.05）。与术前比较,A、B组出麻醉恢复室时非术后认知功能障碍组患儿血浆谷氨酸、γ-氨基丁酸含量略有升高,差异均无统计学意义（P>0.05）,术后认知功能障碍组患儿血浆谷氨酸、γ-氨基丁酸含量均显著升高（P<0.05）。A、B组幼儿出麻醉恢复室时血浆谷氨酸、γ-氨基丁酸含量升高均,与其术后3 d发生术后认知功能障碍正相关（P<0.05）。结论 血浆谷氨酸、γ-氨基丁酸含量可间接反映麻醉后幼儿发生不同程度的脑损伤,可作为幼儿术后早期发生术后认知功能障碍的检测指标。     

Potentiation of inhibitory amino acid receptors-mediated responses by lanthanum in rat sacral dorsal commissural neurons

Lanthanum is one of rare earth cations with extremely active chemical property and has been reported to influence neuronal transmitter systems. To date, little attention has been directed towards the sacral dorsal commissural nucleus (SDCN), which serves as a relay of sensory information from the pelvic viscera in the spinal cord. Therefore, the effect of lanthanum on the inhibitory neurotransmitter γ-aminobutyric acid (GABA) and glycine (Gly) responses in neurons acutely dissociated from the rat SDCN was investigated using the nystatin-perforated patch-recording configuration under voltage-clamp conditions. At a holding potential of − 40 mV, La³⁺ reversibly potentiated GABA (3 μM)-activated currents (I_GABA) in a concentration-dependent manner over the concentration range of 10 μM to 30 mM, with the EC₅₀ value of 67.3 ± 16.4 μM. Similarly, La³⁺ reversibly potentiated glycine (10 μM)-activated currents (I_Gly) in a concentration-dependent manner over the concentration range of 1 μM to 1 mM, with the EC₅₀ value of 52.3 ± 10.9 μM. The effects of La³⁺ on I_GABA and I_Gly were voltage-independent. Moreover, both of the potentiations were not use-dependent and were overcome by increasing the concentration of agonist. Our results indicate that La³⁺ potentiates the inhibitory amino acid receptors-mediated responses in SDCN, which may reduce the transmission of the pelvic visceral information. The information provided by this work may help to elucidate the mechanisms and effects of lanthanum on brain functions.     

Dual effects of pentobarbital on rat sacral dorsal commissural neurons in vitro

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GABAA and GABAB receptors in the nucleus accumbens shell differentially modulate dopamine and acetylcholine receptor-mediated turning behaviour

The ability of GABA_A and GABA_B receptors in the shell of the nucleus accumbens to modulate distinct types of turning behaviour was investigated in freely moving rats, using the unilateral injection technique. The GABA_A receptor agonist muscimol and the GABA_A receptor antagonist bicuculline did not produce turning behaviour; the same holds for the GABA_B agonist baclofen and the GABA_B antagonist 2-hydroxysaclofen. A mixture of the dopamine D₁ receptor agonist SKF 38393 and the dopamine D_2/3 receptor agonist quinpirole has been found to elicit contraversive pivoting, when injected into the shell. This pivoting was dose-dependently inhibited by muscimol, and the inhibitory effect of muscimol was antagonised by bicuculline. Pivoting was also dose-dependently inhibited by baclofen; however, 2-hydroxysaclofen did not antagonise the inhibitory effect. The acetylcholine receptor agonist carbachol has been found to elicit contraversive circling, when injected into the shell. This carbachol-induced circling was inhibited by baclofen, and 2-hydroxysaclofen antagonised the inhibitory effect. Carbachol-induced circling was also partially inhibited by muscimol; however, the inhibitory effect of muscimol was not antagonised by bicuculline. It is concluded that mesolimbic GABA_A receptors exert an inhibitory control on dopamine-dependent pivoting that can be elicited from the shell of the nucleus accumbens, and that GABA_B receptors exert an inhibitory control on acetylcholine-dependent circling that can be elicited from the shell of the nucleus accumbens. This data extends the earlier reported findings that the neurochemical substrate in the shell of the nucleus accumbens that mediates dopamine-dependent pivoting is fundamentally different from the shell substrate that mediates acetylcholine-dependent circling.     

Involvement of GABA and opioid peptide receptors in sevoflurane-induced antinociception in rat spinal cord

AIM: The spinal cord is pivotal in immobility induced by volatile anesthetics because the anesthetics depress the activity of motor neurons in the spinal cord. The aim of this study was to observe the effects of sevoflurane on pain processing at the spinal level. METHODS: The firing of the gastrocnemius muscle was evoked by electrical stimulation to the ipsilateral hindpaw in rats. The nociceptive C response of electromyography (EMG) was selected to study. The GABAA receptor antagonist bicuculline (0.1 mg/kg) and opioid receptor antagonist naloxone (0.4 mg/kg) were administered intravenously, either in the presence or in the absence of 1.0% sevoflurane. RESULTS: In rats with transected spinal cord, sevoflurane produced a profound reduction in the C response in a dose- and time-dependent manner. In the presence of 1.0% sevoflurane, the C responses were increased after injections of bicuculline and naloxone. CONCLUSION: Sevoflurane is a volatile anesthetic that acts directly on the spinal cord to suppress the nociceptive reflex. The sevoflurane-induced suppression of the C response is antagonized by either bicuculline or naloxone. The results suggest that spinal GABAA receptors and opioid peptide receptors are involved in the sevoflurane-induced suppression of spinal nociception.     

5-HT potentiation of the GABAA response in the rat sacral dorsal commissural neurones

1. The modulatory effect of 5-hydroxytryptamine (5-HT) on the γ-aminobutyric acid_A (GABA_A) response was investigated in the neurones freshly dissociated from the rat sacral dorsal commissural nucleus (SDCN) using the nystatin perforated patch recording configuration under the voltage-clamp conditions.
2. 5-HT potentiated GABA-induced Cl⁻ current (I_GABA) without affecting the reversal potential of I_GABA and the apparent affinity of GABA to its receptor.
3. α-Methyl-5-HT mimicked the potentiation effect of 5-HT on I_GABA while ketanserine blocked it. 1-Oleoyl-2-acetyl-glycerol (OAG) potentiated I_GABA, and the effect of 5-HT on I_GABA was occluded by OAG pretreatment. In the presence of chelerythrine, 5-HT failed to potentiate I_GABA, suggesting that protein kinase C (PKC) is involved in the pathway through which the activation of the 5-HT₂ receptor potentiates the I_GABA.
4. The facilitatory effect of 5-HT on I_GABA remained in the presence of BAPTA-AM. LiCl also had no effect on 5-HT-induced potentiation of I_GABA.
5. H-89, genistein, okadaic acid and pervanadate all had no effects on 5-HT potentiation of I_GABA. Pertussis toxin treatment for 6–8 h did not block the facilitatory effect of 5-HT on I_GABA.
6. The present results show that GABA_A receptor in the rat SDCN could be modulated in situ by 5-HT, one of the major transmitters involved in the supraspinal control of nociception, and that the phosphorylation of GABA_A receptor by PKC may be sufficient to support such modulation. The results also strongly support the hypothesis that the cotransmission by 5-HT and GABA has an important role in the spinal cord.

     

Activation of dopamine D4 receptors modulates [3H]GABA release in slices of the rat thalamic reticular nucleus

The thalamic reticular nucleus (nRt) is innervated by dopaminergic projections from the sustantia nigra compacta (SNc) and is rich in dopamine D4 receptors, however, the functional effects of dopamine on this structure are unknown. We examined whether the D1 receptor agonist SKF 38393, or the D2 class receptor agonist quinpirole, modify depolarization evoked Ca(2+)-dependent [3H]GABA release. SKF 38393 was without effects, whereas quinpirole inhibited [3H]GABA release with an IC50 of 81 +/- 33 nM. Dose-dependence determinations of agonists (quinpirole and PD 168, 077) and antagonists (L-745,870, U-101958, clozapine and raclopride) with different affinities for different D2 class subtype receptors showed that a D4 receptor mediates quinpirole inhibition. We used methylphenidate, an agent that acts by increasing interstitial dopamine, to determine whether endogenous dopamine modulates [3H]GABA release. Methylphenidate inhibited [3H]GABA release showing that the nRt contains sufficient endogenous dopamine to activate D4 receptors. This inhibition was completely reversed by selectively blocking D4 receptors with L-745,870 or U-101958 indicating that the catecholamine receptors that modulate GABA release are D4 receptors. Given the importance of the nRt in the control of attention, sensory processing and the generation of rhythmic activity during slow wave sleep, it is possible that abnormal nRt function may generate some of the manifestations of the disorders of dopaminergic transmission.     

Antagonism of baclofen-induced depression of whole-cell synaptic currents in spinal dorsal horn neurones by the potent GABAB antagonist CGP55845

The potencies of two GABA_B receptor antagonists P-[3-aminopropyl]-P-diethoxymethyl-phosphinic acid (CGP35348) and the novel compound 3-N[1-(S)-(3,4-dichlorophenyl)ethyl]amino-2-(S)-hydroxypropyl-P-benzyl-phosphinic acid (CGP55845) have been compared in an in vitro spinal cord preparation. They have been tested as antagonists of baclofen-induced depression of EPSCs of patch-clamped dorsal horn neurones following electrical stimulation of dorsal roots. Mean EC₅₀ values for the depressant action of baclofen were increased by 50- and 140-fold respectively in the presence of CGP35348 (200 μM) (n = 5) and CGP55845 (100 nM) (n = 4). This potency of CGP55845 is > 1000-fold higher than that reported previously for other GABA_B receptor antagonists.