We have newly developed a system that allows monitoring of the intensity of fluorescent signals from deep brains of rats transgenically modified to express enhanced green fluorescent protein (eGFP) via an optical fiber. One terminal of the optical fiber was connected to a blue semiconductor laser oscillator/green fluorescence detector. The other terminal was inserted into the vicinity of the eGFP-expressing neurons. Since the optical fiber was vulnerable to twisting stresses caused by animal movement
we also developed a cage in which the floor automatically turns
in response to the turning of the rat’s head. This relieved the twisting stress on the optical fiber. The system then enabled real-time monitoring of fluorescence in awake and unrestrained rats over many hours. Using this system
we could continuously monitor eGFP-expression in arginine vasopressin-eGFP transgenic rats. Moreover
we observed an increase of eGFP-expression in the paraventricular nucleus under salt-loading conditions. We then performed in vivo imaging of eGFP-expressing GnRH neurons in the hypothalamus
via a bundle consisting of 3000 thin optical fibers. With the combination of the optical fiber bundle connection to the fluorescence microscope
and the special cage system
we were able to capture and retain images of eGFP-expressing neurons from free-moving rats. We believe that our newly developed method for monitoring and imaging eGFP-expression in deep brain neurons will be useful for analysis of neuronal functions in awake and unrestrained animals for long durations.
