Bardóczi Zsuzsanna, Pál Balázs, Kőszeghy Áron, Wilheim Tamás, Watanabe Masahiko, Záborszky László, Liposits Zsolt, Kalló Imre
Laboratory of Endocrine Neurobiology, Institute of Experimental Medicine, HAS, 1083, Budapest, Hungary.
Semmelweis University, School of PH.D. Studies, 1085, Budapest, Hungary.
J Neurosci. 2017 Sep 27;37(39):9534-9549. doi: 10.1523/JNEUROSCI.3348-16.2017. Epub 2017 Sep 5.
The basal forebrain (BF) receives afferents from brainstem ascending pathways, which has been implicated first by Moruzzi and Magoun (1949) to induce forebrain activation and cortical arousal/waking behavior; however, it is very little known about how brainstem inhibitory inputs affect cholinergic functions. In the current study, glycine, a major inhibitory neurotransmitter of brainstem neurons, and gliotransmitter of local glial cells, was tested for potential interaction with BF cholinergic (BFC) neurons in male mice. In the BF, glycine receptor α subunit-immunoreactive (IR) sites were localized in choline acetyltransferase (ChAT)-IR neurons. The effect of glycine on BFC neurons was demonstrated by bicuculline-resistant, strychnine-sensitive spontaneous IPSCs (sIPSCs; 0.81 ± 0.25 × 10 Hz) recorded in whole-cell conditions. Potential neuronal as well as glial sources of glycine were indicated in the extracellular space of cholinergic neurons by glycine transporter type 1 (GLYT1)- and GLYT2-IR processes found in apposition to ChAT-IR cells. Ultrastructural analyses identified synapses of GLYT2-positive axon terminals on ChAT-IR neurons, as well as GLYT1-positive astroglial processes, which were localized in the vicinity of synapses of ChAT-IR neurons. The brainstem raphe magnus was determined to be a major source of glycinergic axons traced retrogradely from the BF. Our results indicate a direct effect of glycine on BFC neurons. Furthermore, the presence of high levels of plasma membrane glycine transporters in the vicinity of cholinergic neurons suggests a tight control of extracellular glycine in the BF. Basal forebrain cholinergic (BFC) neurons receive various activating inputs from specific brainstem areas and channel this information to the cortex via multiple projections. So far, very little is known about inhibitory brainstem afferents to the BF. The current study established glycine as a major regulator of BFC neurons by (1) identifying glycinergic neurons in the brainstem projecting to the BF, (2) showing glycine receptor α subunit-immunoreactive (IR) sites in choline acetyltransferase (ChAT)-IR neurons, (3) demonstrating glycine transporter type 2 (GLYT2)-positive axon terminals synapsing on ChAT-IR neurons, and (4) localizing GLYT1-positive astroglial processes in the vicinity of synapses of ChAT-IR neurons. The effect of glycine on BFC neurons was demonstrated by bicuculline-resistant, strychnine-sensitive spontaneous IPSCs recorded in whole-cell conditions.
基底前脑（BF）接收来自脑干上行通路的传入神经，莫鲁齐和马贡在1949年首次指出这与诱导前脑激活及皮质觉醒/清醒行为有关；然而，关于脑干抑制性输入如何影响胆碱能功能却知之甚少。在本研究中，对雄性小鼠的基底前脑胆碱能（BFC）神经元进行了测试，以检测甘氨酸（一种脑干神经元的主要抑制性神经递质和局部神经胶质细胞的胶质递质）与其之间的潜在相互作用。在基底前脑中，甘氨酸受体α亚基免疫反应性（IR）位点定位于胆碱乙酰转移酶（ChAT）免疫反应性神经元中。在全细胞条件下记录到的荷包牡丹碱抗性、士的宁敏感的自发性抑制性突触后电流（sIPSCs；0.81±0.25×10Hz）证明了甘氨酸对BFC神经元的作用。在胆碱能神经元的细胞外空间中，通过与ChAT免疫反应性细胞相邻的1型甘氨酸转运体（GLYT1）和GLYT2免疫反应性过程，表明了甘氨酸的潜在神经元及神经胶质来源。超微结构分析确定了GLYT2阳性轴突终末与ChAT免疫反应性神经元之间的突触，以及GLYT1阳性星形胶质细胞过程，它们定位于ChAT免疫反应性神经元突触附近。经逆行追踪确定脑干中缝大核是来自基底前脑的甘氨酸能轴突的主要来源。我们的结果表明甘氨酸对BFC神经元有直接作用。此外，胆碱能神经元附近高水平的质膜甘氨酸转运体的存在表明基底前脑中细胞外甘氨酸受到严格控制。基底前脑胆碱能（BFC）神经元从特定脑干区域接收各种激活输入，并通过多个投射将这些信息传递到皮质。到目前为止，关于脑干对基底前脑的抑制性传入神经知之甚少。本研究通过以下几点确定甘氨酸是BFC神经元的主要调节因子：（1）确定投射到基底前脑的脑干中的甘氨酸能神经元；（2）显示胆碱乙酰转移酶（ChAT）免疫反应性神经元中的甘氨酸受体α亚基免疫反应性（IR）位点；（3）证明GLYT2阳性轴突终末与ChAT免疫反应性神经元形成突触；（4）将GLYT1阳性星形胶质细胞过程定位于ChAT免疫反应性神经元突触附近。在全细胞条件下记录到的荷包牡丹碱抗性、士的宁敏感的自发性抑制性突触后电流证明了甘氨酸对BFC神经元的作用。
