Frequency-Dependent Cannabinoid Receptor-Independent Modulation of Glycine Receptors by Endocannabinoid 2-AG

Endocannabinoids are known as retrograde messengers, being released from the postsynaptic neuron and acting on specific presynaptic G-protein-coupled cannabinoid (CB) receptors to decrease neurotransmitter release. Also, at physiologically relevant concentrations cannabinoids can directly modulate the function of voltage-gated and receptor-operated ion channels. Using patch-clamp recording we analyzed the consequences of the direct action of an endocannabinoid, 2-arachidonoylglycerol (2-AG), on the functional properties of glycine receptor channels (GlyRs) and ionic currents in glycinergic synapses. At physiologically relevant concentrations (0.1–1 μM), 2-AG directly affected the functions of recombinant homomeric α1H GlyR: it inhibited peak amplitude and dramatically enhanced desensitization. The action of 2-AG on GlyR-mediated currents developed rapidly, within ∼300 ms. Addition of 1 μM 2-AG strongly facilitated the depression of glycine-induced currents during repetitive (4–10 Hz) application of short (2 ms duration) pulses of glycine to outside-out patches. In brainstem slices from CB1 receptor knockout mice, 2-AG significantly decreased the extent of facilitation of synaptic currents in hypoglossal motoneurons during repetitive (10–20 Hz) stimulation. These observations suggest that endocannabinoids can modulate postsynaptic metaplasticity of glycinergic synaptic currents in a CB1 receptor-independent manner

Recombinant ionotropic glutamate receptors: functional distinctions imparted by different subunits

Molecular cloning and functional analysis of recombinantly expressed ionotropic glutamate receptors (GluR) have revealed a large variety of GluR subunits that could be grouped into three families: AMPA, kainate and NMDA receptors. Different kinetic and ion permeation properties of the channels formed from GluR subunits were obtained using human embryonic kidney 293 cells as an expression system and fast agonist application combined with whole-cell current recording. Common structural elements responsible for ion permeation through recombinant GluR channels are identified by point mutation analysis. These data may provide a guideline for the identification of the molecular composition of native GluR channels on the basis of both their functional properties and the expression pattern of their subunits

Calcium permeability of glutamate-gated channels in the central nervous system

Molecular cloning of ionotropic glutamate receptors and the development of new measurement techniques have significantly advanced our understanding of the molecular mechanisms controlling ligand-mediated entry of Ca2+ into neurons of the mammalian CNS. Recent studies have demonstrated that various types of glutamate receptors expressed in different nerve cells are permeable to Ca2+ to variable extents, depending on the structural peculiarities of the subunits and their composition in a particular cell. This diversity provides a regulable pathway for Ca2+ entry during synaptic transmission. The fractional contribution of this Ca2+ to the total synaptic current might be a substantial means of elevating the intracellular Ca2+ concentration over a wide temporal range

Functional Analysis of NMDAR Subunit Components in Postsynaptic Currents of Identified Cells and Synapses in Brain Slices

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Block of native Ca2+ permeable AMPA receptors in rat brain by intracellular polyamines generates double rectification

1. The influence of intracellular factors on current rectification of different subtypes of native alpha-amino-3-hydroxy-5-methyl-4- isoxazoleproprionate receptors (AMPARs) was studied in rat brain slices by combining fast application of glutamate with patch pipette perfusion. 2. The peak current-voltage (I-V) relation of the AMPARs expressed in Bergmann glial cells of cerebellum and dentate gyrus (DG) basket cells of hippocampus was weakly rectifying in outside-out patches and nystatin-perforated vesicles, but showed a doubly rectifying shape with a region of reduced slope between 0 and +40 mV in nucleated patches. The I-V relation of AMPARs expressed in hippocampal CA3 pyramidal neurones was linear in all recording configurations. 3. Intracellular application of 25 microM spermine, a naturally occurring polyamine, blocked outward currents in outside-out patches from Bergmann glial cells and DG basket cells in a voltage-dependent manner, generating I-V relations with a doubly rectifying shape which were similar to those recorded in nucleated patches. AMPARs in CA3 pyramidal cell patches were unaffected by 25 microM spermine. 4. The half-maximal blocking concentration of spermine at +40 mV was 0.3 microM in Bergmann glial cell patches and 1.5 microM in DG basket cell patches, whereas it was much higher (>> 100 microM) for CA3 pyramidal cell patches. Spermidine also affected current rectification, but with lower affinity. The block of outward current by polyamines following voltage jumps developed within < 0.5 ms. 5. We conclude that current rectification, rather than being an intrinsic property of the Ca(2+)- permeable AMPAR channel, is generated by polyamine block

Determinants of Ca2+ permeability in both TM1 and TM2 of high affinity kainate receptor channels: diversity by RNA editing

GluR6, a subunit of high affinity kainate receptor channels in the mammalian CNS, carries a glutamine (Q) or arginine (R) residue in a critical position (Q/R site) of the putative channel-forming segment TM2. One form, GluR6(Q), is encoded by the GluR6 gene; the other, GluR6(R), is generated by RNA editing. Further analysis of cloned GluR6 cDNA revealed that two additional positions, located in transmembrane segment TM1, are diversified by RNA editing to generate either isoleucine (I) or valine (V) in one and tyrosine (Y) or cysteine (C) in the other TM1 position. In GluR6 channels, in contrast with AMPA receptor channels, the presence of Q in the TM2 Q/R site determines channels with low Ca2+ permeability, whereas an R determines a higher Ca2+ permeability if TM1 is fully edited. In the TM1 unedited form of GluR6, Ca2+ permeability is less dependent on the presence of either Q or R in TM2. Thus Ca2+ permeability of kainate receptor channels can vary, depending on editing of both TM1 and TM2