Role of Ca^<2+> channel in the function of gonadotropin-releasing hormone surge generator in rats

Ca^2通道在大鼠促性腺激素释放激素浪涌发生器功能中的作用

• 批准号: 11670075
• 负责人: TANAKA Fukuko
• 金额: $ 2.3万
• 依托单位: Yokohama City University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11670075/
• 关键词: Rat; GnRH surge generator; LH surge; TTX; Ca^<2+> channel; GABA neurons; Calcium imaging; Na^+ channel; Ca2+チャネル; 視索前野; Na+チャネル; GnRHサージジェネレー; Ca^<2+>チャネルター; Na^+チャネル

1. Effects of tetrodotoxin (TTX) injected into the medial preoptic area (MPO) on electrical stimulation-induced luteinizing hormone (LH) surge in pentobarbital (PB)-anesthetized proestrous ratsRats exhibiting regular 4-days estrous cycle were injected with 35 mg/kg PB, which is known to block the LH surge, on the day of proestrus at 13 : 45. Thereafter, rats were injected with 1, 2, or 5 μM TTX dissolved in 0.5 μl saline into the MPO and they received an electrical stimulation for 30 min, which is known to induce the LH surge in PB-blocked proestrous rats. Some rats were injected with saline as a control. The result showed that injection of 1, 2, or 5 μM TTX into the MPO blocked the electrical stimulation-induced LH surge in PB-blocked proestrous rats.2. Effects of TTX injected into the MPO on the spontaneous LH surge in proestrous ratsRats exhibiting regular 4-days estrous cycle were injected into the bilateral MPO with 5 μM TTX dissolved in 1.5 μl each on the day of proestrus at 14 : … More 30. Some rats were injected with saline as control. Bilateral injection of TTX into the MPO completely blocked spontaneous LH surge.3. Effects of Ca^<2+> channel blocker injected into the MPO on the spontaneous LH surge in proestrous ratsRats exhibiting regular 4-days estrous cycle were injected into the bilateral MPO with 5 μM nifedipine, a L-type Ca^<2+> channel blocker, dissolved in 1.5 ml each side on the day of proestrus at 14 : 30. Some rats were injected with saline as control. Bilateral injection of nifedipine into the MPO completely blocked spontaneous LH surge.4. Macro Ca^<2+> imaging analysis in MPO slices obtained from proestrous rats in association with the activity of GnRH surge generatorOn the morning of proestrus, rats were killed between 10 : 00 and 11 : 00 and 200-μm-thick PDA slices were cut with a brain slicer. After 1 h of incubation in a standard artificial cerebrospinal fluid (aCSF) containing 10 mM Calcium green-1, POA slices were perfused with aCSF at a flow rate of 500 μl/min. Changes in [Ca^<2+>]I were determined by a macro imaging analysis. We could not see spontaneous fluctuation of [Ca^<2+>]I but 100 μM bicuculline induced an increase in [Ca^<2+>]I and GnRH secretion in POA slices. In a separate experiment, we found that infusion of 100 μM bicuculline significantly increased GnRH secretion but infusion of 100 μM naloxone was without effect.In summary, results suggests that the activation of TTX-sensitive voltage-dependent Na^+ channel and L-type Ca^<2+> channel are important for the activity of GnRH surge generator. Further, as we hypothesized, it is suggested that a disinhibition of inhibitory GABA tone on GnRH neurons in the POA elicits a surge of GnRH secretion but that a disinhibition of inhibitory opioid tone does not. Less

1.在戊巴比妥（PB）麻醉的动情前期大鼠，视前内侧区（MPO）注射河豚毒素（TTX）对电刺激诱发的促黄体生成激素（LH）峰的影响在动情前期的当天13：45注射阻断LH峰的PB 35 mg/kg。然后，将溶于0.5 μl生理盐水中的1、2或5 μM TTX注射到大鼠的MPO中，并接受电刺激30 min，这是已知的在PB阻断的动情前期大鼠中诱导LH峰。一些大鼠注射生理盐水作为对照。结果表明，在PB阻断的动情前期大鼠，MPO内注射1、2、5 μM TTX均能阻断电刺激诱导的LH峰. MPO内注射TTX对动情前期大鼠自发性LH峰的影响动情前期14：00将5 μM TTX溶于1.5 μl/只，注入动情前期大鼠双侧MPO内。 ...更多信息 30.部分大鼠注射生理盐水作为对照。双侧MPO注射TTX完全阻断自发性LH峰。3. MPO内注射Ca^<2+>通道阻滞剂对动情前期大鼠自发性LH峰的影响在动情前期的当天14：30，将L型Ca^<2+>通道阻滞剂硝苯地平（5 μM）每侧1.5ml注射到动情前期大鼠的双侧MPO内。部分大鼠注射生理盐水作为对照。双侧髓过氧化物酶内注射硝苯地平完全阻断自发性LH峰.动情前期大鼠脑片MPO钙离子宏观显像分析与GnRH峰发生器活性的相关性动情前期早晨10：00 ~ 11：00处死大鼠，用脑切片机切取200 μ m厚的PDA脑片。在含有10 mM钙绿色-1的标准人工脑脊液（aCSF）中孵育1小时后，以500 μl/min的流速用aCSF灌注POA切片。通过宏观成像分析测定[Ca^<2+>]I的变化。我们没有看到[Ca^<2+>]I的自发波动，但100 μM荷包牡丹碱可诱导POA切片中[Ca^<2+>]I和GnRH分泌增加。在另一项实验中，我们发现100 μM荷包牡丹碱可显著增加GnRH的分泌，而100 μM纳洛酮则无此作用。总之，结果表明，TTX敏感的电压依赖性Na^+通道和L型Ca^<2+>通道的激活对GnRH浪涌发生器的活性起重要作用。此外，正如我们假设的那样，这表明对POA中GnRH神经元的抑制性GABA张力的解除抑制会引起GnRH分泌的激增，但抑制性阿片张力的解除抑制不会。少

项目成果

Funabashi T, Mitsushima D, Nakamura JT, Shinohara K. Suyama K, Kimura F: "Gonadotropin-releasing hormone (GnRH) surge generator in female rats"Progress in Brain Research (special issue for 2nd international symposium on the comparative biology of GnRH). (

Funabashi T，Mitsushima D，Nakamura JT，Shinohara K. Suyama K，Kimura F：“雌性大鼠促性腺激素释放激素（GnRH）浪涌发生器”脑研究进展（第二届GnRH比较生物学国际研讨会特刊）

舩橋利也, 貴邑冨久子: "内側視索前野GnRHニューロンと性周期"Clinical Neuroscience. 19. 1122-1124 (2001)

Toshiya Funahashi、Tomiko Takamura：“内侧视前区 GnRH 神经元和性周期”《临床神经科学》19. 1122-1124 (2001)。

Funabashi T, Shinohara K, Kimura F: "Neuronal control circuit for the gonadotropin-releasing hormone surge in rats. In Neuroplasticity, development, and steroid hormone action (Handa RJ, Hayashi S, Terasawa E, Kawata M, eds)"CRC Press. Florida. 169-176 (2

Funabashi T、Shinohara K、Kimura F：“大鼠促性腺激素释放激素激增的神经控制回路。神经可塑性、发育和类固醇激素作用（Handa RJ、Hayashi S、Terasawa E、Kawata M，编辑）”CRC Press

Funabashi T, Aiba S, Sano A, Shinohara K, Kimura F: "Intracerebroventricular injection of arginine-vasopressin V1 receptor antagonist attenuates the surge of luteinizing hormone and prolactin secretion in proestrous rats"Neuroscience Letters. 260. 37-40 (

Funabashi T、Aiba S、Sano A、Shinohara K、Kimura F：“脑室内注射精氨酸-加压素 V1 受体拮抗剂可减弱发情前大鼠黄体生成素和催乳素分泌的激增”《神经科学快报》。

Funabashi T, Aiba S, Sano A, Shinohara K. Kimura F: "Intracerebroventricular injection of arginine-vasopressin Vl receptor antagonist attenuates the surge of luteinizing hormone and prolactin secretion in proestrous rats"Neuroscience Letters. 260. 37-40 (

Funabashi T、Aiba S、Sano A、Shinohara K. Kimura F：“脑室内注射精氨酸-加压素 VI 受体拮抗剂可减弱发情前大鼠黄体生成素和催乳素分泌的激增”《神经科学快报》。