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Gap junctions have been postulated to exist between the axons of excitatory cortical neurons based on electrophysiological, modeling, and dye-coupling data. Here, we provide ultrastructural evidence for axoaxonic gap junctions in dentate granule cells. Using combined confocal laser scanning microscopy, thin-section transmission electron microscopy, and grid-mapped freeze-fracture replica immunogold labeling, 10 close appositions revealing axoaxonic gap junctions ([approximately equal to] 30-70 nm in diameter) were found between pairs of mossy fiber axons ([approximately equal to] 100-200 nm in diameter) in the stratum lucidum of the CA3b field of the rat ventral hippocampus, and one axonal gap junction ([approximately equal to] 100 connexons) was found on a mossy fiber axon in the CA3c field of the rat dorsal hippocampus. Immunogold labeling with two sizes of gold beads revealed that connexin36 was present in that axonal gap junction. These ultrastructural data support computer modeling and in vitro electrophysiological data suggesting that axoaxonic gap junctions play an important role in the generation of very fast (>70 Hz) network oscillations and in the hypersynchronous electrical activity of epilepsy. axoaxonic | connexin | electrical synapse | epilepsy | synchronization

Cone photoreceptors mediate daylight vision in vertebrates. Changes in neurotransmitter release at cone synapses encode visual information and is subject to precise control by negative feedback from enigmatic horizontal cells. However, the mechanisms that orchestrate this modulation are poorly understood due to a virtually unknown landscape of molecular players. Here, we report a molecular player operating selectively at cone synapses that modulates effects of horizontal cells on synaptic release. Using an unbiased proteomic screen, we identified an adhesion GPCR Latrophilin3 (LPHN3) in horizontal cell dendrites that engages in transsynaptic control of cones. We detected and characterized a prominent splice isoform of LPHN3 that excludes a element with inhibitory influence on transsynaptic interactions. A gain-of-function mouse model specifically routing LPHN3 splicing to this isoform but not knockout of LPHN3 diminished CaV1.4 calcium channel activity profoundly disrupted synaptic release by cones and resulted in synaptic transmission deficits. These findings offer molecular insight into horizontal cell modulation on cone synaptic function and more broadly demonstrate the importance of alternative splicing in adhesion GPCRs for their physiological function. [ABSTRACT FROM AUTHOR]

Establishment of functional synaptic connections in a selective manner is essential for nervous system operation. In mammalian retinas, rod and cone photoreceptors form selective synaptic connections with different classes of bipolar cells (BCs) to propagate light signals. While there has been progress in elucidating rod wiring, molecular mechanisms used by cones to establish functional synapses with BCs have remained unknown. Using an unbiased proteomic strategy in cone-dominant species, we identified the cell-adhesion molecule ELFN2 to be pivotal for the functional wiring of cones with the ON type of BC. It is selectively expressed in cones and transsynaptically recruits the key neurotransmitter receptor mGluR6 in ON-BCs to enable synaptic transmission. Remarkably, ELFN2 in cone terminals functions in synergy with a related adhesion molecule, ELFN1, and their concerted interplay during development specifies selective wiring and transmission of cone signals. These findings identify a synaptic connectivity mechanism of cones and illustrate how interplay between adhesion molecules and postsynaptic transmitter receptors orchestrates functional synaptic specification in a neural circuit. [ABSTRACT FROM AUTHOR]

Exposure of cultured primary neurons to preformed α-synuclein fibrils (PFFs) leads to the recruitment of endogenous α-synuclein and its templated conversion into fibrillar phosphorylated α-synuclein (pα-synF) aggregates resembling those involved in Parkinson's disease (PD) pathogenesis. Pα-synF was described previously as inclusions morphologically similar to Lewy bodies and Lewy neurites in PD patients. We discovered the existence of a conformationally distinct, nonfibrillar, phosphorylated α-syn species that we named "pα-syn*." We uniquely describe the existence of pα-syn* in PFF-seeded primary neurons, mice brains, and PD patients' brains. Through immunofluorescence and pharmacological manipulation we showed that pα-syn* results from incomplete autophagic degradation of pα-synF. Pα-synF was decorated with autophagic markers, but pα-syn* was not. Western blots revealed that pα-syn* was N- and C-terminally trimmed, resulting in a 12.5-kDa fragment and a SDS-resistant dimer. After lysosomal release, pα-syn* aggregates associated with mitochondria, inducing mitochondrial membrane depolarization, cytochrome C release, and mitochondrial fragmentation visualized by confocal and stimulated emission depletion nanoscopy. Pα-syn* recruited phosphorylated acetyl-CoA carboxylase 1 (ACC1) with which it remarkably colocalized. ACC1 phosphorylation indicates low ATP levels, AMPK activation, and oxidative stress and induces mitochondrial fragmentation via reduced lipoylation. Pα-syn* also colocalized with BiP, a master regulator of the unfolded protein response and a resident protein of mitochondria-associated endoplasmic reticulum membranes that are sites of mitochondrial fission and mitophagy. Pα-syn* aggregates were found in Parkin-positive mitophagic vacuoles and imaged by electron microscopy. Collectively, we showed that pα-syn* induces mitochondrial toxicity and fission, energetic stress, and mitophagy, implicating pα-syn* as a key neurotoxic α-syn species and a therapeutic target. [ABSTRACT FROM AUTHOR]

The time course of signaling via heterotrimeric G proteins is controlled through their activation by G-protein coupled receptors and deactivation through the action of GTPase accelerating proteins (GAPs). Here we identify RGS7 and RGS11 as the key GAPs in the mGluR6 pathway of retinal rod ON bipolar cells that set the sensitivity and time course of light-evoked responses. We showed using electroretinography and single cell recordings that the elimination of RGS7 did not influence dark-adapted light-evoked responses, but the concurrent elimination of RGS11 severely reduced their magnitude and dramatically slowed the onset of the response. In RGS7/RGS11 double-knockout mice, light-evoked responses in rod ON bipolar cells were only observed during persistent activation of rod photoreceptors that saturate rods. These observations are consistent with persistently high G-protein activity in rod ON bipolar cell dendrites caused by the absence of the dominant GAP, biasing TRPM1 channels to the closed state. [ABSTRACT FROM AUTHOR]

Cone photoreceptors mediate daylight vision in vertebrates. Changes in neurotransmitter release at cone synapses encode visual information and is subject to precise control by negative feedback from enigmatic horizontal cells. However, the mechanisms that orchestrate this modulation are poorly understood due to a virtually unknown landscape of molecular players. Here, we report a molecular player operating selectively at cone synapses that modulates effects of horizontal cells on synaptic release. Using an unbiased proteomic screen, we identified an adhesion GPCR Latrophilin3 (LPHN3) in horizontal cell dendrites that engages in transsynaptic control of cones. We detected and characterized a prominent splice isoform of LPHN3 that excludes a element with inhibitory influence on transsynaptic interactions. A gain-of-function mouse model specifically routing LPHN3 splicing to this isoform but not knockout of LPHN3 diminished Ca V 1.4 calcium channel activity profoundly disrupted synaptic release by cones and resulted in synaptic transmission deficits. These findings offer molecular insight into horizontal cell modulation on cone synaptic function and more broadly demonstrate the importance of alternative splicing in adhesion GPCRs for their physiological function., Competing Interests: The authors declare no competing interest.

Establishment of functional synaptic connections in a selective manner is essential for nervous system operation. In mammalian retinas, rod and cone photoreceptors form selective synaptic connections with different classes of bipolar cells (BCs) to propagate light signals. While there has been progress in elucidating rod wiring, molecular mechanisms used by cones to establish functional synapses with BCs have remained unknown. Using an unbiased proteomic strategy in cone-dominant species, we identified the cell-adhesion molecule ELFN2 to be pivotal for the functional wiring of cones with the ON type of BC. It is selectively expressed in cones and transsynaptically recruits the key neurotransmitter receptor mGluR6 in ON-BCs to enable synaptic transmission. Remarkably, ELFN2 in cone terminals functions in synergy with a related adhesion molecule, ELFN1, and their concerted interplay during development specifies selective wiring and transmission of cone signals. These findings identify a synaptic connectivity mechanism of cones and illustrate how interplay between adhesion molecules and postsynaptic transmitter receptors orchestrates functional synaptic specification in a neural circuit., Competing Interests: The authors declare no competing interest.

The time course of signaling via heterotrimeric G proteins is controlled through their activation by G-protein coupled receptors and deactivation through the action of GTPase accelerating proteins (GAPs). Here we identify RGS7 and RGS11 as the key GAPs in the mGluR6 pathway of retinal rod ON bipolar cells that set the sensitivity and time course of light-evoked responses. We showed using electroretinography and single cell recordings that the elimination of RGS7 did not influence dark-adapted light-evoked responses, but the concurrent elimination of RGS11 severely reduced their magnitude and dramatically slowed the onset of the response. In RGS7/RGS11 double-knockout mice, light-evoked responses in rod ON bipolar cells were only observed during persistent activation of rod photoreceptors that saturate rods. These observations are consistent with persistently high G-protein activity in rod ON bipolar cell dendrites caused by the absence of the dominant GAP, biasing TRPM1 channels to the closed state.