The death of photoreceptor cells caused by retinal degenerative diseases often results in a complete loss of retinal responses to light. We explore the feasibility of converting inner retinal neurons to photosensitive cells as a possible strategy for imparting light sensitivity to retinas lacking rods and cones. Using delivery by an adeno-associated viral vector, here, we show that long-term expression of a microbial-type rhodopsin, channelrhodopsin-2 (ChR2), can be achieved in rodent inner retinal neurons in vivo. Furthermore, we demonstrate that expression of ChR2 in surviving inner retinal neurons of a mouse with photoreceptor degeneration can restore the ability of the retina to encode light signals and transmit the light signals to the visual cortex. Thus, expression of microbial-type channelrhodopsins, such as ChR2, in surviving inner retinal neurons is a potential strategy for the restoration of vision after rod and cone degeneration.

Two groups of retinal cone bipolar cells (CBCs) in rats were found to express voltage-gated Na + channels. The axon terminals of the first group stratify in sublamina 2 of the inner plexiform layer (IPL) and partially overlap with the OFF-cholinergic band. This group was identified as type 3 CBCs. The axon terminals of the second group stratify in sublamina 3 of the IPL, slightly distal to the ONcholinergic band. Cells of this second group resemble type 5 CBCs. In addition, we observed another group of ON-type CBCs with terminal stratification similar to that of the second group. However, this latter group did not show any Na + current, instead exhibiting a large hyperpolarization-activated cyclic nucleotide-gated cation current, suggesting the existence of two subclasses of physiologically distinct type 5 CBCs. Both groups of Na + -expressing bipolar cells were capable of generating a rapid TTX sensitive action potential as revealed by current injection. Multiple spike-like potentials were also observed in some of these cells. Results of this study provide valuable insights into the function of voltage-gated Na + channels of retinal bipolar cells in retinal processing.

Journal of Physiology Physiology in PressRBCs in the rat. We first assessed the spatial distribution of glycine receptors on RBCs by direct patch-clamp recordings of isolated presynaptic terminals and by focal puffing of glycine in retinal slices. We found that glycine receptors are highly concentrated at the axon terminals of RBCs. The pharmacological and biophysical properties of glycine receptors located in the axon terminal and somatic/dendritic regions are similar. Furthermore, we present evidence for the existence of glycinergic synaptic inputs onto the axon terminals of RBCs and show that activation of glycine receptors could effectively suppress depolarization-evoked calcium influx into the axon terminals. METHODS Dissociation of bipolar cellsBipolar cells were dissociated from Long-Evans rats ≤4 weeks of age as described previously Pan, 2000). All animal handling procedures were approved by the Institutional Animal Care Committee at Wayne State University, and were in accord with the NIH Guide for the Care and Use of Laboratory Animals. In brief, animals were deeply anaesthetized with CO 2 and killed by decapitation. Retinas were removed and placed in a Hanks' solution (mM): NaCl, 138; NaHCO 3 , 1; Na 2 HPO 4 , 0.3; KCl, 5; KH 2 PO 4 , 0.3; CaCl 2 , 1.25 or 2.5; MgSO 4 , 0.5; MgCl 2 , 0.5; Hepes-NaOH, 5; glucose, 22.2; with phenol red, 0.001 % v/v; pH 7.2. The retinas were incubated for ~40-50 min at 34-37°C in an enzymatic solution that consisted of Hanks' solution (described above), supplemented with DL-cysteine, 0.2 mg ml _1 ; bovine serum albumin, 0.2 mg ml _1 ; and papain, ~2 u ml _1 . After several rinses in Hanks' solution, the retinas were mechanically dissociated by gentle trituration with a glass pipette. The dissociated cells were plated onto culture dishes in normal or Ca 2+ -free Hanks' solution. Cells were kept at room temperature and used for recordings within 5 h. The use of Ca 2+ -free Hanks' solution for cell culture was found to help maintain healthier axon terminals of bipolar cells. The Ca 2+ -free Hanks' solution is the same as the Hanks' solution described above except for the omission of Ca 2+ ions.

In axon-bearing neurons, action potentials conventionally initiate at the axon initial segment (AIS) and are important for neuron excitability and cell-to-cell communication. However in axonless neurons, spike origin has remained unclear. Here we report in the axonless spiking AII amacrine cell of the mouse retina a dendritic process sharing organizational and functional similarities with the AIS. This process was revealed through viral-mediated expression of channelrhodopsin-2-GFP (ChR2-GFP) with the AIS-targeting motif of sodium channels (NavII-III). The AII-processes showed clustering of voltage-gated Na+ channel 1.1 (Nav1.1) as well as AIS markers ankyrin-G and neurofascin. Furthermore, NavII-III targeting disrupted Nav1.1 clustering in the AII-process which drastically decreased Na+ current and abolished the ability of the AII amacrine cell to generate spiking. Our findings indicate that despite lacking an axon, spiking in the axonless neuron can originate at a specialized AIS-like process.

Retinal bipolar cells comprise multiple subtypes that are well known for the diversity of their physiological properties. We investigated the properties and functional roles of the hyperpolarization-activated currents in mammalian retinal bipolar cells using whole cell patch-clamp recording techniques. We report that bipolar cells express inwardly rectifying K+ currents ( IKir) in addition to the hyperpolarization-activated cationic currents ( Ih) previously reported. Furthermore, these two currents are differentially expressed among different subtypes of bipolar cells. One group of cone bipolar cells in particular displayed mainly IKir. A second group of cone bipolar cells displayed both currents but with a much larger Ih. Rod bipolar cells, on the other hand, showed primarily Ih. Moreover, we showed that IKir and Ih differentially influence the voltage responses of bipolar cells: Ih facilitates and/or accelerates the membrane potential rebound, whereas IKir counteracts or prevents such rebound. The findings of the expression of IKir and the differential expression of Ih and IKir in bipolar cells may provide new insights into an understanding of the physiological properties of bipolar cells.