(A) Schematics of the NINscope with dimensions in mm. Two 10 by 10 mm HDI printed circuit boards (PCBs), one for interfacing with the data acquisition box (Interface PCB), the other containing the CMOS imaging sensor (Sensor PCB), are stacked and mounted in a 3D printed enclosure. Excitation light from an LED is collimated with a half ball lens, passes through an excitation filter and is reflected by the dichroic mirror onto the specimen. The emitted fluorescence is collected through the GRIN objective lens, and passes the dichroic and a plano-convex lens, which focuses an image onto the CMOS sensor. An emission filter is glued onto the plano-convex lens with optical bonding glue. (B) KiCad renders of the custom-built interface and sensor PCBs with top and bottom views. The interface PCB contains an inertial measurement unit (IMU) for measuring head acceleration and orientation, three LED drivers including one for optogenetic (strobe) control and two for excitation LEDs (one used), as well as a red tracking LED, the serializer, and the IO expander. The sensor PCB contains the PYTHON480 CMOS sensor, clock oscillator and a power sequencer, which provides the image sensor with the necessary voltages as well as their timing and sequence. (C) Photograph of NINscope with coax and strain cables, excitation LED and optogenetic LED cables. (D) Custom-built implantable LED probe for optogenetic stimulation that is connected to NINscope using the optogenetic LED cable. (E) The UCLA Miniscope DAQ card v3.2 was used with minor modifications including a 256 kB x 8-bit I2C EEPROM (STMicroelectronics) and a wired connection bridging general purpose input/output 2 (GPIO2) with test point 4 (TP4). The serial peripheral interface (SPI) signals: master output slave input (MOSI), serial clk (SCK) and slave select (SSN) are connected to GPIO0, GPIO1 and GPIO3 through jumpers.