Monitoring neuronal activity in vivo is critical to understanding the physiological or pathological functions of the brain. Two-photon Ca²⁺ imaging in vivo using a cranial window and specific neuronal labeling enables real-time, in situ, and long-term imaging of the living brain. Here, we constructed a recombinant rabies virus containing the Ca²⁺ indicator GCaMP6s along with the fluorescent protein DsRed2 as a baseline reference to ensure GCaMP6s signal reliability. This functional tracer was applied to retrogradely label specific V1–thalamus circuits and detect spontaneous Ca²⁺ activity in the dendrites of V1 corticothalamic neurons by in vivo two-photon Ca²⁺ imaging. Notably, we were able to record single-spine spontaneous Ca²⁺ activity in specific circuits. Distinct spontaneous Ca²⁺ dynamics in dendrites of V1 corticothalamic neurons were found for different V1–thalamus circuits. Our method can be applied to monitor Ca²⁺ dynamics in specific input circuits in vivo, and contribute to functional studies of defined neural circuits and the dissection of functional circuit connections.

中文翻译：

---

狂犬病病毒标记的V1皮层丘脑神经元树突状体的体内双光子钙成像。

体内监测神经元活动对于了解大脑的生理或病理功能至关重要。使用颅窗和特定神经元标记在体内进行双光子Ca ²⁺成像，可以对活体大脑进行实时，原位和长期成像。在这里，我们构建了包含Ca ²⁺指示剂GCaMP6s和荧光蛋白DsRed2作为基线参考的重组狂犬病毒，以确保GCaMP6s信号的可靠性。该功能示踪剂用于体内逆行标记特定的V1-丘脑回路，并通过体内检测V1皮质丘脑神经元树突中的自发Ca ²⁺活性双光子Ca ²⁺成像。值得注意的是，我们能够在特定电路中记录单根脊柱自发性Ca ²⁺活性。在不同的V1-丘脑回路中，发现V1皮层丘脑神经元的树突中有明显的自发Ca ²⁺动力学。我们的方法可用于监测体内特定输入回路中Ca ^2+的动力学，并有助于定义神经回路的功能研究和功能电路连接的解剖。