The stream of visual information sent from photoreceptors to second-order bipolar cells is intercepted by laterally interacting horizontal cells that generate feedback to optimize and improve the efficiency of signal transmission. The mechanisms underlying the regulation of graded photoreceptor synaptic output in this nonspiking network have remained elusive. Here, we analyze with patch clamp recording the novel mechanisms by which horizontal cells control pH in the synaptic cleft to modulate photoreceptor neurotransmitter release. First, we show that mammalian horizontal cells respond to their own GABA release and that the results of this autaptic action affect cone voltage-gated Ca 2+ channel (Ca V channel) gating through changes in pH. As a proof-of-principle, we demonstrate that chemogenetic manipulation of horizontal cells with exogenous anion channel expression mimics GABA-mediated cone Ca V channel inhibition. Activation of these GABA receptor anion channels can depolarize horizontal cells and increase cleft acidity via Na + /H + exchanger (NHE) proton extrusion, which results in inhibition of cone Ca V channels. This action is effectively counteracted when horizontal cells are sufficiently hyperpolarized by increased GABA receptor (GABAR)-mediated HCO 3 − efflux, alkalinizing the cleft and disinhibiting cone Ca V channels. This demonstrates how hybrid actions of GABA operate in parallel to effect voltage-dependent pH changes, a novel mechanism for regulating synaptic output.

Summary Compartmentalized signaling in dendritic subdomains is critical for the function of many central neurons. In the retina, individual dendritic sectors of a starburst amacrine cell (SAC) are preferentially activated by different directions of linear motion, indicating limited signal propagation between the sectors. However, the mechanism that regulates this propagation is poorly understood. Here, we find that metabotropic glutamate receptor 2 (mGluR2) signaling, which acts on voltage-gated calcium channels in SACs, selectively restricts cross-sector signal propagation in SACs, but does not affect local dendritic computation within individual sectors. mGluR2 signaling ensures sufficient electrotonic isolation of dendritic sectors to prevent their depolarization during non-preferred motion, yet enables controlled multicompartmental signal integration that enhances responses to preferred motion. Furthermore, mGluR2-mediated dendritic compartmentalization in SACs is important for the functional output of direction-selective ganglion cells (DSGCs). Therefore, our results directly link modulation of dendritic compartmentalization to circuit-level encoding of motion direction in the retina.

The cellular mechanisms underlying feedback signaling from horizontal cells to photoreceptors, which are important for the formation of receptive field surrounds of early visual neurons, remain unsettled. Mammalian horizontal cells express a complement of synaptic proteins that are necessary and sufficient for calcium-dependent exocytosis of inhibitory neurotransmitters at their contacts with photoreceptor terminals, suggesting that they are capable of releasing GABA via vesicular release. To test whether horizontal cell vesicular release is involved in feedback signaling, we perturbed inhibitory neurotransmission in these cells by targeted deletion of the vesicular GABA transporter (VGAT), the protein responsible for the uptake of inhibitory transmitter by synaptic vesicles. To manipulate horizontal cells selectively, an iCre mouse line with Cre recombinase expression controlled by connexin57 (Cx57) regulatory elements was generated. In Cx57-iCre mouse retina, only horizontal cells expressed Cre protein, and its expression occurred in all retinal regions. After crossing with a VGAT flox/flox mouse line, VGAT was selectively eliminated from horizontal cells, which was confirmed immunohistochemically. Voltage-gated ion channel currents in horizontal cells of Cx57-VGAT Ϫ/Ϫ mice were the same as Cx57-VGAT ϩ/ϩ controls, as were the cell responses to the ionotropic glutamate receptor agonist kainate, but the response to the GABA A receptor agonist muscimol in Cx57-VGAT Ϫ/Ϫ mice was larger. In contrast, the feedback inhibition of photoreceptor calcium channels, which in control animals is induced by horizontal cell depolarization, was completely absent in Cx57-VGAT Ϫ/Ϫ mice. The results suggest that vesicular release of GABA from horizontal cells is required for feedback inhibition of photoreceptors. Key words: Ca channels; Cx57-iCre; GABA receptors; inhibitory feedback; retinal horizontal cells; synaptic vesicles Significance StatementFeedback inhibition by horizontal cells regulates the photoreceptor calcium channels responsible for the release of the neurotransmitter glutamate. This feedback inhibition contributes to the formation of the receptive field surrounds of early visual neurons, which is important for contrast sensitivity and color opponency in vision. The cellular mechanisms underlying feedback inhibition are not fully understood. One disputed mechanism for mammalian horizontal cell inhibitory output involves the vesicular release of the inhibitory neurotransmitter GABA. By selectively deleting the transporter that normally loads GABA into vesicles in horizontal cells, we show that horizontal cells lose their ability to modulate photoreceptor calcium signals. These findings implicate a role for the vesicular release of GABA in mediating inhibitory feedback.

Food and water are rewarding in part because they satisfy our internal needs1,2. Dopaminergic neurons in the ventral tegmental area (VTA) are activated by gustatory rewards3–5, but how animals learn to associate these oral cues with the delayed physiological effects of ingestion is unknown. Here we show that individual dopaminergic neurons in the VTA respond to detection of nutrients or water at specific stages of ingestion. A major subset of dopaminergic neurons tracks changes in systemic hydration that occur tens of minutes after thirsty mice drink water, whereas different dopaminergic neurons respond to nutrients in the gastrointestinal tract. We show that information about fluid balance is transmitted to the VTA by a hypothalamic pathway and then re-routed to downstream circuits that track the oral, gastrointestinal and post-absorptive stages of ingestion. To investigate the function of these signals, we used a paradigm in which a fluid’s oral and post-absorptive effects can be independently manipulated and temporally separated. We show that mice rapidly learn to prefer one fluid over another based solely on its rehydrating ability and that this post-ingestive learning is prevented if dopaminergic neurons in the VTA are selectively silenced after consumption. These findings reveal that the midbrain dopamine system contains subsystems that track different modalities and stages of ingestion, on timescales from seconds to tens of minutes, and that this information is used to drive learning about the consequences of ingestion.