Multidisciplinary Institute of Cell Biology (IMBICE), National Council of Science and Technology (CONICET), Buenos Aires Comision of Science (CIC) and La Plata University (UNLP), La Plata, Argentina.
J Physiol. 2018 Nov;596(22):5415-5428. doi: 10.1113/JP276256. Epub 2018 Oct 14.
Presynaptic Ca 2 voltage-gated calcium channels link action potentials arriving at the presynaptic terminal to neurotransmitter release. Hence, their regulation is essential to fine tune brain circuitry. Ca 2 channels are highly sensitive to G protein-coupled receptor (GPCR) modulation. Our previous data indicated that growth hormone secretagogue receptor (GHSR) constitutive activity impairs Ca 2 channels by decreasing their surface density. We present compelling support for the impact of Ca 2.2 channel inhibition by agonist-independent GHSR activity exclusively on GABA release in hippocampal cultures. We found that this selectivity arises from a high reliance of GABA release on Ca 2.2 rather than on Ca 2.1 channels. Our data provide new information on the effects of the ghrelin-GHSR system on synaptic transmission, suggesting a putative physiological role of the constitutive signalling of a GPCR that is expressed at high levels in brain areas with restricted access to its natural agonist.
Growth hormone secretagogue receptor (GHSR) displays high constitutive activity, independent of its endogenous ligand, ghrelin. Unlike ghrelin-induced GHSR activity, the physiological role of GHSR constitutive activity and the mechanisms that underlie GHSR neuronal modulation remain elusive. We previously demonstrated that GHSR constitutive activity modulates presynaptic Ca 2 voltage-gated calcium channels. Here we postulate that GHSR constitutive activity-mediated modulation of Ca 2 channels could be relevant in the hippocampus since this brain area has high GHSR expression but restricted access to ghrelin. We performed whole-cell patch-clamp in hippocampal primary cultures from E16- to E18-day-old C57BL6 wild-type and GHSR-deficient mice after manipulating GHSR expression with lentiviral transduction. We found that GHSR constitutive activity impairs Ca 2.1 and Ca 2.2 native calcium currents and that Ca 2.2 basal impairment leads to a decrease in GABA but not glutamate release. We postulated that this selective effect is related to a higher Ca 2.2 over Ca 2.1 contribution to GABA release (∼40% for Ca 2.2 in wild-type vs. ∼20% in wild-type GHSR-overexpressing cultures). This effect of GHSR constitutive activity is conserved in hippocampal brain slices, where GHSR constitutive activity reduces local GABAergic transmission of the granule cell layer (intra-granule cell inhibitory postsynaptic current (IPSC) size ∼-67 pA in wild-type vs. ∼-100 pA in GHSR-deficient mice), whereas the glutamatergic output from the dentate gyrus to CA3 remains unchanged. In summary, we found that GHSR constitutive activity impairs IPSCs both in hippocampal primary cultures and in brain slices through a Ca 2-dependent mechanism without affecting glutamatergic transmission.
突触前 Ca2+电压门控钙通道将到达突触前末端的动作电位与神经递质释放联系起来。因此，它们的调节对于微调大脑回路至关重要。Ca2+通道对 G 蛋白偶联受体（GPCR）的调节非常敏感。我们之前的数据表明，生长激素促分泌素受体（GHSR）的组成性活性通过降低其表面密度来损害 Ca2+通道。我们提供了令人信服的证据，证明激动剂非依赖性 GHSR 活性对 Ca2.2 通道的抑制作用仅对海马培养物中的 GABA 释放有影响。我们发现，这种选择性源自 GABA 释放对 Ca2.2 的高度依赖，而不是对 Ca2.1 通道的依赖。我们的数据提供了关于促胃液素 - GHSR 系统对突触传递影响的新信息，表明在大脑区域中表达高水平的 GPCR 的组成性信号传导可能具有生理作用，而该区域对其天然激动剂的进入受到限制。
生长激素促分泌素受体（GHSR）表现出高组成性活性，而不依赖其内源性配体，即 ghrelin。与 ghrelin 诱导的 GHSR 活性不同，GHSR 组成性活性的生理作用和 GHSR 神经元调节的机制仍不清楚。我们之前证明 GHSR 组成性活性调节突触前 Ca2+电压门控钙通道。在这里，我们假设 GHSR 组成性活性介导的 Ca2+通道调节可能与海马体有关，因为该脑区具有高 GHSR 表达，但对 ghrelin 的进入受到限制。我们使用慢病毒转导来操纵 GHSR 表达后，在来自 E16 至 E18 天龄 C57BL6 野生型和 GHSR 缺陷型小鼠的海马体原代培养物中进行全细胞膜片钳记录。我们发现 GHSR 组成性活性损害 Ca2.1 和 Ca2.2 天然钙电流，并且 Ca2.2 基础损害导致 GABA 但不是谷氨酸释放减少。我们假设这种选择性效应与 GABA 释放中 Ca2.2 对 Ca2.1 的贡献较高有关（野生型中约为 40%，而野生型中 GHSR 过表达培养物中约为 20%）。GHSR 组成性活性的这种作用在海马脑片中得以保留，其中 GHSR 组成性活性降低颗粒细胞层的局部 GABA 能传递（野生型中的内颗粒细胞抑制性突触后电流（IPSC）大小约为-67 pA，而 GHSR 缺陷型小鼠中的约为-100 pA），而来自齿状回的谷氨酸能输出到 CA3 保持不变。总之，我们发现 GHSR 组成性活性通过 Ca2+依赖性机制损害海马体原代培养物和脑片中的 IPSC，而不影响谷氨酸能传递。
