, 2005 and Gunaydin et al., 2010) to toxicity (Gradinaru et al., 2008, Gradinaru et al., 2010 and Zhao et al., 2008) to challenges linked to light delivery in vivo (Aravanis et al., 2007 and Adamantidis et al., 2007). A long process of tool engineering and substantial development of enabling technologies was required over the next several years. The key properties of these microbial optogenetic tools relate to the ecology of their original host organisms, XAV-939 which respond to the environment using seven-transmembrane proteins encoded by the type I

class of opsin gene (Yizhar et al., 2011b). Type I opsins are protein products of microbial opsin genes and are termed rhodopsins when bound to retinal. However, in typical heterologous expression experiments the precise composition of retinoid-bound states is uncharacterized.

Therefore in the setting of neuroscience application, the tools are conservatively referred to as opsins (a more accurate and convenient shorthand for common use, since only “opsin” correctly applies to the genes as well as to the protein products). These proteins are distinguished from their PLX-4720 molecular weight mammalian (type II) counterparts, in that they are single-component light-sensing systems; the two operations—light sensing and ion conductance—are carried out by the same protein. The first identified, and still by far the best studied, type I protein is the haloarchaeal proton pump bacteriorhodopsin (BR; Figure 1A; Oesterhelt and Stoeckenius, 1971, Oesterhelt and Stoeckenius, 1973 and Racker and Stoeckenius, 1974). Under low-oxygen conditions, BR is highly expressed in haloarchaeal membranes and serves as part of an alternative energy-production system, pumping protons from the cytoplasm to the extracellular medium to generate a proton-motive force to drive ATP synthesis (Racker and Stoeckenius, 1974 and Michel and Oesterhelt, 1976). These light-gated proton pumps have since

also been found in a wide range of marine proteobacteria as well as in other kingdoms of life, where they employ similar photocycles (Béjà et al., 2001 and Váró et al., 2003) and have been hypothesized to play diverse roles in cellular Idoxuridine physiology (Fuhrman et al., 2008). A second class of microbial opsin genes encodes halorhodopsins (Figure 1A). Halorhodopsin (HR) is a light-activated chloride pump first discovered in archaebacteria (Matsuno-Yagi and Mukohata, 1977). The operating principles of halorhodopsin (HR) are similar to those of BR (Essen, 2002), with the two main differences being that halorhodopsin pumps chloride ions and its direction of transport is from the extracellular to the intracellular space. Specific amino acid residues have been shown to underlie the differences between BR and HR in directionality and preferred cargo ion (Sasaki et al., 1995).