MiR-51Regulates GABAergic Synaptic Function By GLO-4in C.elegan S

miRNAs is a class of endogenous non-coding RNAs, with the size of about20to25nucleotides. They are often conserved among the species during evolution. It isreported that miRNAs are particularly enriched in the nervous system, and are closelyrelated to many physiological processes of the nervous system, including neuronaldifferentiation, neurite outgrowth, synaptic plasticity. In many nervous diseases, theabnormal expression of different types of miRNAs can be detected. Some evidencesshowed that miRNAs are related to the synaptic functions. mir-1was known to playan important regulatory role in neuromuscular connections and postsynaptic function.mir-134is expressed in the synapse dendritic cells in hippocampal neurons, which is anegative regulator of postsynaptic excitatory synaptic transmission by controlling thesize of the postsynaptic dendritic spines. However, it is still unclear if miRNAs areinvolved in GABAergic synaptic function. The neural structure of C. elegans is verysimple, which is easy to be observed. The genes related to neural functions in C.elegans have high homology to those in mammals. C. elegans contains a variety ofclassic neurotransmitters in mammals, such as acetylcholine, γ-aminobutyric acid,glutamate, dopamine, serotonin, and neuropeptides. These neurotransmitters areinvolved in the process of neuronal storage, synthesis and metabolism with the similarmechanism of mammals. In addition, half of the C. elegans miRNAs have thesequence homology to miRNAs in humans. Therefore, C. elegans are very suitable forresearching the mechanism of miRNAs in GABAeric synapses functions.Our previous study has found that mir-51affected GABAergic synaptic function.When treated with GABA antagonists PTZ, the mir-51mutants appeared thephenotype of hypersensitivity to PTZ. Moreover, we found that glo-4is a target geneof mir-51by RT-PCR and dual luciferase reporter assay. Therefore, we intend to knowwhether miR-51can regulate GABAergic synapses function by its target gene glo-4.First, we constructed mir-51; glo-4double mutants. And, by PTZ-inducedconvulsions analysis, we found that mir-51can affect GABA synapse functions byGLO-4. Then, we use GFP fluorescence to label the GABAergic synapses ofwild-type N2, mir-51mutants, glo-4mutants and mir-51; glo-4double mutants, respectively. SNB-1::GFP (juIs1) is used to label the presynaptic membrane as thesynaptic binding protein, whose density of the fluorescent dots mean the synapticdensity. We found that the density of SNB-1puncta was reduced in mir-51mutants.However, the reduction of SNB-1puncta density was rescued by loss of function inglo-4. These results suggested that mir-51regulated the GABAergic synaptogenesisby GLO-4. UNC-49::GFP (oxIs22) is utilized to mark GABAAreceptor abundance onthe postsynaptic membrane. Our results showed that mir-51mutants reduced thedensity of UNC-49puncta. And the decreasing of UNC-49puncta density induced bymir-51mutants was recovered by deletion of glo-4. These indicated that mir-51regulated the abundance of GABAAreceptor by GLO-4. PUNC-47::GFP (oxIs12) is usedto check the axonal growth in GABAergic synapses. We found that defective of bothmir-51and glo-4affected the axonal growth in GABAergic synapses. This suggestedthat mir-51is associated with the axonal growth in GABAergic synapses independenton GLO-4. These results indicated that mir-51regulated synaptogenesis and GABAAreceptor abundance at GABAergic synapses by GLO-4. This will lay a theoreticalfoundation for the study of the mechanism that miRNAs are involved in the regulationof GABAergic synapses, and provide new clues for the therapy of theGABAergic related diseases.