MTA-SZTE Research Group for Cortical Microcircuits of the Hungarian Academy of Sciences, Department of Physiology, Anatomy and Neuroscience, University of Szeged, Közép fasor 52, Szeged, H-6726, Hungary.
1st Department of Internal Medicine, University of Szeged, Szeged, Hungary.
Diabetologia. 2019 Apr;62(4):717-725. doi: 10.1007/s00125-018-4803-z. Epub 2019 Jan 12.
AIMS/HYPOTHESIS: Glucagon-like peptide 1 (GLP-1) receptors are expressed by pancreatic beta cells and GLP-1 receptor signalling promotes insulin secretion. GLP-1 receptor agonists have neural effects and are therapeutically promising for mild cognitive impairment and Alzheimer's disease. Our previous results showed that insulin is released by neurogliaform neurons in the cerebral cortex, but the expression of GLP-1 receptors on insulin-producing neocortical neurons has not been tested. In this study, we aimed to determine whether GLP-1 receptors are present in insulin-containing neurons.
We harvested the cytoplasm of electrophysiologically and anatomically identified neurogliaform interneurons during patch-clamp recordings performed in slices of rat neocortex. Using single-cell digital PCR, we determined copy numbers of Glp1r mRNA and other key genes in neurogliaform cells harvested in conditions corresponding to hypoglycaemia (0.5 mmol/l glucose) and hyperglycaemia (10 mmol/l glucose). In addition, we performed whole-cell patch-clamp recordings on neurogliaform cells to test the effects of GLP-1 receptor agonists for functional validation of single-cell digital PCR results.
Single-cell digital PCR revealed GLP-1 receptor expression in neurogliaform cells and showed that copy numbers of mRNA of the Glp1r gene in hyperglycaemia exceeded those in hypoglycaemia by 9.6 times (p < 0.008). Moreover, single-cell digital PCR confirmed co-expression of Glp1r and Ins2 mRNA in neurogliaform cells. Functional expression of GLP-1 receptors was confirmed with whole-cell patch-clamp electrophysiology, showing a reversible effect of GLP-1 on neurogliaform cells. This effect was prevented by pre-treatment with the GLP-1 receptor-specific antagonist exendin-3(9-39) and was absent in hypoglycaemia. In addition, single-cell digital PCR of neurogliaform cells revealed that the expression of transcription factors (Pdx1, Isl1, Mafb) are important in beta cell development.
CONCLUSIONS/INTERPRETATION: Our results provide evidence for the functional expression of GLP-1 receptors in neurons known to release insulin in the cerebral cortex. Hyperglycaemia increases the expression of GLP-1 receptors in neurogliaform cells, suggesting that endogenous incretins and therapeutic GLP-1 receptor agonists might have effects on these neurons, similar to those in pancreatic beta cells.
目的/假设：胰高血糖素样肽 1 (GLP-1) 受体存在于胰岛β细胞中，GLP-1 受体信号转导可促进胰岛素分泌。GLP-1 受体激动剂具有神经作用，对轻度认知障碍和阿尔茨海默病具有治疗前景。我们之前的研究结果表明，胰岛素由大脑皮层中的神经胶质形神经元释放，但胰岛素产生的新皮层神经元上是否存在 GLP-1 受体尚未得到测试。在这项研究中，我们旨在确定 GLP-1 受体是否存在于含有胰岛素的神经元中。
我们在大鼠新皮层切片中进行膜片钳记录时，收获电生理和解剖学上鉴定的神经胶质形中间神经元的细胞质。使用单细胞数字 PCR，我们确定了在对应于低血糖（0.5mmol/l 葡萄糖）和高血糖（10mmol/l 葡萄糖）条件下收获的神经胶质形细胞中 Glp1r mRNA 和其他关键基因的拷贝数。此外，我们对神经胶质形细胞进行全细胞膜片钳记录，以测试 GLP-1 受体激动剂的作用，以验证单细胞数字 PCR 结果的功能。
单细胞数字 PCR 显示 GLP-1 受体在神经胶质形细胞中表达，并表明高血糖时 Glp1r 基因的 mRNA 拷贝数比低血糖时高 9.6 倍（p<0.008）。此外，单细胞数字 PCR 证实神经胶质形细胞中 Glp1r 和 Ins2 mRNA 的共表达。全细胞膜片钳电生理学证实 GLP-1 受体的功能表达，表明 GLP-1 对神经胶质形细胞具有可逆作用。这种作用可被 GLP-1 受体特异性拮抗剂 exendin-3(9-39) 预处理所阻止，并且在低血糖时不存在。此外，神经胶质形细胞的单细胞数字 PCR 显示，转录因子（Pdx1、Isl1、Mafb）的表达对β细胞发育很重要。
结论/解释：我们的研究结果为大脑皮层中已知释放胰岛素的神经元中 GLP-1 受体的功能表达提供了证据。高血糖增加神经胶质形细胞中 GLP-1 受体的表达，提示内源性肠促胰岛素和治疗性 GLP-1 受体激动剂可能对这些神经元产生与胰岛β细胞类似的作用。
