Liprin-␣ proteins are adaptors that interact with the receptor protein tyrosine phosphatase leukocyte common antigen-related (LAR) and other synaptic proteins to promote synaptic partner selection and active zone assembly. Liprin-␤ proteins bind to and share homology with Liprin-␣ proteins, but their functions at the synapse are unknown. The Drosophila genome encodes single Liprin-␣ and Liprin-␤ homologs, as well as a third related protein that we named Liprin-␥. We show that both Liprin-␤ and Liprin-␥ physically interact with Liprin-␣ and that Liprin-␥ also binds to LAR. Liprin-␣ mutations have been shown to disrupt synaptic target layer selection by R7 photoreceptors and to reduce the size of larval neuromuscular synapses. We have generated null mutations in Liprin-␤ and Liprin-␥ to investigate their role in these processes. We find that, although Liprin-␣ mutant R7 axons terminate before reaching the correct target layer, Liprin-␤ mutant R7 axons grow beyond their target layer. Larval neuromuscular junction size is reduced in both Liprin-␣ and Liprin-␤ mutants, and further reduced in double mutants, suggesting independent functions for these Liprins. Genetic interactions demonstrate that both Liprin proteins act through the exchange factor Trio to promote stable target selection by R7 photoreceptor axons and growth of neuromuscular synapses. Photoreceptor and neuromuscular synapses develop normally in Liprin-␥ mutants; however, removing Liprin-␥ improves R7 targeting in Liprin-␣ mutants, and restores normal neuromuscular junction size to Liprin-␤ mutants, suggesting that Liprin-␥ counteracts the functions of the other two Liprins. We propose that context-dependent interactions between the three Liprins modulate their functions in synapse formation.

One unique subtype of retinal ganglion cell is the direction selective (DS) cell, which responds vigorously to stimulus movement in a preferred direction, but weakly to movement in the opposite or null direction. Here we show that the application of the GABA receptor blocker picrotoxin unmasks a robust excitatory OFF response in ON DS ganglion cells. Similar to the characteristic ON response of ON DS cells, the masked OFF response is also direction selective, but its preferred direction is opposite to that of the ON component. Given that the OFF response is unmasked with picrotoxin, its direction selectivity cannot be generated by a GABAergic mechanism. Alternatively, we find that the direction selectivity of the OFF response is blocked by cholinergic drugs, suggesting that acetylcholine release from presynaptic starburst amacrine cells is crucial for its generation. Finally, we find that the OFF response is abolished by application of a gap junction blocker, suggesting that it arises from electrical synapses between ON DS and polyaxonal amacrine cells. Our results suggest a novel role for gap junctions in mixing excitatory ON and OFF signals at the ganglion cell level. We propose that OFF inputs to ON DS cells are normally masked by a GABAergic inhibition, but are unmasked under certain stimulus conditions to mediate optokinetic signals in the brain.

We have examined the role of neighbor relationships between cholinergic amacrine cells upon their positioning and dendritic field size by producing partial ablations of this population of cells during early development. We first determined the effectiveness of l-glutamate as an excitotoxin for ablating cholinergic amacrine cells in the developing mouse retina. Subcutaneous injections (4 mg0g) made on P-3 and thereafter were found to produce a near-complete elimination, while injections at P-2 were ineffective. Lower doses on P-3 produced only partial reductions, and were subsequently used to examine the effect of partial ablation upon mosaic organization and dendritic growth of the remaining cells. Four different Voronoi-based measures of mosaic geometry were examined in l-glutamate-treated and normal (saline-treated) retinas. Partial depletions of around 40% produced cholinergic mosaics that, when scaled for density, approximated the mosaic geometry of the normal retina. Separate comparisons simulating a 40% random deletion of the normal retina produced mosaics that were no different from those experimentally depleted retinas. Consequently, no evidence was found for positional regulation in the absence of normal neighbor relationships. Single cells in the ganglion cell layer were intracellularly filled with Lucifer Yellow to examine the morphology and dendritic field extent following partial ablation of the cholinergic amacrine cells. No discernable effect was found on their starburst morphology, and total dendritic field area, number of primary dendrites, and branch frequency were not significantly different. Cholinergic amacrine cells normally increase their dendritic field area after P-3 in excess of retinal expansion; despite this, the present results show that this growth is not controlled by the density of neighboring processes.

Non-technical summary An organizing principle of the visual system is the segregation of ON and OFF responses into parallel streams to signal light increment and decrement. This segregation begins in the retina where the output ganglion cells can be divided into ON and OFF subtypes based on their responses to light. Here we show that blockade of GABAergic inhibition reveals opposite polarity responses in ganglion cells whereby OFF cells display ON responses and ON cells display OFF responses. This crossover excitation was found in both the rabbit and mouse, indicating that it is a common synaptic mechanism in the mammalian retina. Overall, these results challenge the idea that light increment and decrement is signalled by different visual pathways. Moreover, our findings suggest that release of inhibition under certain light conditions can enable single ganglion cells to carry both ON and OFF signals, thereby allowing additional information to be propagated across the limited bandwidth of the optic nerve.Abstract A fundamental organizing feature of the visual system is the segregation of ON and OFF responses into parallel streams to signal light increment and decrement. However, we found that blockade of GABAergic inhibition unmasks robust ON responses in OFF α-ganglion cells (α-GCs). These ON responses had the same centre-mediated structure as the classic OFF responses of OFF α-GCs, but were abolished following disruption of the ON pathway with L-AP4. Experiments showed that both GABA A and GABA C receptors are involved in the masking inhibition of this ON response, located at presynaptic inhibitory synapses on bipolar cell axon terminals and possibly amacrine cell dendrites. Since the dendrites of OFF α-GCs are not positioned to receive excitatory inputs from ON bipolar cell axon terminals in sublamina-b of the inner plexiform layer (IPL), we investigated the possibility that gap junction-mediated electrical synapses made with neighbouring amacrine cells form the avenue for reception of ON signals. We found that the application of gap junction blockers eliminated the unmasked ON responses in OFF α-GCs, while the classic OFF responses remained. Furthermore, we found that amacrine cells coupled to OFF α-GCs display processes in both sublaminae of the IPL, thus forming a plausible substrate for the reception and delivery of ON signals to OFF α-GCs. Finally, using a multielectrode array, we found that masked ON and OFF signals are displayed by over one-third of ganglion cells in the rabbit and mouse retinas, suggesting that masked crossover excitation is a widespread phenomenon in the inner mammalian retina.

Mindfulness training has been shown to improve attention and change the underlying brain substrates in adults. Most mindfulness training programs involve a myriad of techniques, and it is difficult to attribute changes to any particular aspect of the program. Here, we created a video game, Tenacity, which models a specific mindfulness technique – focused attention on one’s breathing – and assessed its potential to train an attentional network in adolescents. A combined analysis of resting state functional connectivity (rs-FC) and diffusion tensor imaging (DTI) yielded convergent results – change in communication within the left fronto-parietal network after two weeks of playing Tenacity compared to a control game. Rs-FC analysis showed greater connectivity between left dorsolateral prefrontal cortex (dlPFC) and left inferior parietal cortex (IPC) in the Tenacity group. Importantly, changes in left dlPFC – IPC rs-FC and changes in structural connectivity of the white matter tract that connects these regions –left superior longitudinal fasiculus (SLF) – were associated with changes in performance on an attention task. Finally, changes in left dlPFC – IPC rs-FC correlated with the change in left SLF structural connectivity as measured by fractional anisotropy (FA) in the Tenacity group only.