The functional principle of the vertebrate brain is often paralleled to a computer: information collected by dedicated devices is processed and integrated by interneuron circuits and leads to output. light-sensory motorneurons and light-sensory interneurons are also present in the brains PX 12 of vertebrates challenging the paradigm that information processing and output circuitry in the central brain is shielded from direct environmental influences. We investigated two groups of nonvisual photopigments VAL- and TMT-Opsins in zebrafish and medaka fish; two teleost species from distinct habitats separated by over 300 million years of evolution. TMT-Opsin subclasses are specifically expressed not only in hypothalamic and thalamic deep brain photoreceptors but also in interneurons and motorneurons with no known photoreceptive function such as the typeXIV interneurons of the fish optic tectum. We further show that TMT-Opsins and Encephalopsin render neuronal cells light-sensitive. TMT-Opsins preferentially respond to blue light relative to rhodopsin with subclass-specific response kinetics. We discovered that co-express with mRNA specifically to neurosecretory cells in the preoptic area/hypothalamus challenging the hypothesis that TMT-Opsin would function as a peripheral light receptor . We PX 12 started our study by an investigation of TMT-Opsin using the vertebrate model organisms zebrafish (hybridization experiments on adult brain sections reveal that are expressed not only in photoreceptive cells of the pineal interneurons in the eye and presumptive CSF-contacting neurosecretory cells. They are also expressed PX 12 in brain nuclei neither connected to light sensation nor neurosecretion such as the dorsal tegmental nucleus the nucleus semicircular torus the facial nerve nucleus the periventricular layer of the tectum and the granular layer of the olfactory bulb. In addition to the evolutionary conservation in expression over at least 300 million years anti-TMTopsin1b staining reveals protein expression in neurons in these brain Parp8 nuclei suggesting that these opsins exhibit functions important for the animals. The neuronal nature of several (cells. Expression of TMT-Opsins does not only render neuronal cells light sensitive in tissue culture but we also show that interneurons at the position of typeXIV interneurons in isolated tectal slices of adult brains show electrophysiological responses to light. Finally we find that a subpopulation of and/or and gene in tissue culture PX 12 in a light-dependent manner . We next tested the relative spectral sensitivity of TMT-Opsins. We adjusted the light intensity using a set of band-pass and neutral density filters in order to activate human rhodopsin equally strong with a blue and a green filter thus calibrating the setup to the rhodopsin absorption maximum of 497 nm ( Figure S4A-B). To minimize well-to-well variations we compared CI changes to color light with a preceding white light stimulus (results of quantification given as relative response (%AUC) Figure S4C). All tested TMT-Opsins (medaka TMT-Opsin 1b 2 and 3a) responded stronger to blue light (λmax 450 nm) compared to green (λmax 528 nm) or red (λmax 605 nm) light than human rhodopsin (Figures 2G and S4C-E). Also the absolute photon number values of the tested wavelengths were highly similar PX 12 (see Figure S4 and Materials and Methods). None of the TMT-Opsins responded to near infrared light (950 nm Figure S5). We also tested the light sensitivity of Encephalopsin. The murine Encephalopsin ortholog has been shown to be specifically expressed in brain interneurons . However no evidence for a photoreceptor function of any Encephalopsin exists so far. With our assay we obtained clear light-dependent responses for medaka Encephalopsin in both Neuro-2A and HEK cells (Figure 2H I). Even though the response PX 12 maximum was lower than for TMT-Opsins the response kinetics exhibited the typical trend seen for opsin activation (compare Figure 2H and Figure S6). Again mutation of Lysine 296 to Alanine abolished the light-dependent response (Figure 2H I) thus providing first evidence that Encephalopsins can function as light receptors. TMT-Opsins Show Specific Expression in Larval and Adult Medaka Fish.