ABSTRACT
In cerebellar cortex, mGlu4 receptors located on parallel fibers play an essential role in normal motor function, but the molecular mechanisms involved are not yet completely understood. Using a strategy combining biochemical and electrophysiological approaches in the rodent cerebellum, we demonstrate that presynaptic mGlu4 receptors control synaptic transmission through an atypical activation of Gαq proteins. First, the Gαq subunit, PLC and PKC signaling proteins present in cerebellar extracts are retained on affinity chromatography columns grafted with different sequences of the cytoplasmic domain of mGlu4 receptor. The i2 loop and the C terminal domain were used as baits, two domains that are known to play a pivotal role in coupling selectivity and efficacy. Second, in situ proximity ligation assays show that native mGlu4 receptors and Gαq subunits are in close physical proximity in cerebellar cortical slices. Finally, electrophysiological experiments demonstrate that the molecular mechanisms underlying mGlu4 receptor-mediated inhibition of transmitter release at cerebellar Parallel Fiber (PF) - Molecular Layer Interneuron (MLI) synapses involves the Gαq-PLC signaling pathway. Taken together, our results provide compelling evidence that, in the rodent cerebellar cortex, mGlu4 receptors act by coupling to the Gαq protein and PLC effector system to reduce glutamate synaptic transmission.
Subject(s)
Cerebellar Cortex/cytology , GTP-Binding Protein alpha Subunits, Gq-G11/metabolism , Receptors, Metabotropic Glutamate/metabolism , Signal Transduction/physiology , Synaptic Transmission/physiology , Animals , Animals, Newborn , Benzopyrans/pharmacology , Cytoplasm/metabolism , Enzyme Activation/drug effects , Enzyme Inhibitors/pharmacology , Excitatory Amino Acid Agents/pharmacology , Excitatory Postsynaptic Potentials/drug effects , Excitatory Postsynaptic Potentials/genetics , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Models, Biological , Nerve Net/drug effects , Propionates/pharmacology , Rats , Rats, Sprague-Dawley , Receptors, Metabotropic Glutamate/genetics , Signal Transduction/drug effects , Synaptic Transmission/drug effects , Synaptic Transmission/geneticsABSTRACT
The total soluble protein content in crude extracts of gonads ofHelix aspersa Müller from different stages of its life cycle was quantified and then resolved into different protein fractions using polyacrylamide gel electrophoresis. It was found that both the protein content and wet weight of the gonads increased during growth but decreased markedly during the copulation and egg-laying phases. Electrophoretic analysis of the extracts from different stages revealed that additional fractions progressively appeared and that in adults, the number and comcentration of the bands changed in response to physiological conditions (activity, copulation, egg-laying or hibernation) so that the maximum number of 27 fractions was observed during the active phase. We have attempted to correlate these biochemical observations with previously published descriptions of histological changes that occur during the development of the gonad.
