GABA_B Receptors Modulate Glutamate Release And Action In Rat Spinal Substantia Gelatinosa

The substantia gelatinosa(SG,lamina II of spinal cord gray matter),mainly composed of interneurons and glia,is pivotal for receiving and regulating the nociceptive information from the peripheral to the central nervous system.γ-Aminobutyric acid type B receptors(GABA_BRs),the metabotropic type of the GABA receptors,are widely expressed in the pre-and postsynaptic structures of the SG neurons.It has been well documented that activation of GABA_BRs by exogenous agonists induces both pre-and postsynaptic inhibition.However,the actions of endogenous GABA via presynaptic GABA_BRs in regulating glutamatergic release,and the postsynaptic GABA_BRs interaction with glutamate,remain largely elusive.The present study identified the mechanism of endogenous GABA in regulating presynaptic glutamate release and postsynaptic excitatory glutamate responses by using whole-cell patch-clamp recordings and holographic photostimulation.Furthermore,we employed Golgi staining to elucidate the morphological characterization of SG neurons and summarize the trend of their dendrites.Taken together,the results give new clues for endogenous GABA in modulating the nociception circuit of the spinal dorsal horn and shed fresh light on the analgesic effects of GABA_Breceptors at the spinal cord level.Objective To investigate the effects of endogenous GABA on glutamatergic synapses through presynaptic GABA_BRs,the interaction between the postsynaptic GABA_BRs and glutamate,and the morphological characteristics of SG neurons.Methods(1)Using patch-clamp recordings with different stimulation strategies and GABA_Breceptor antagonist CGP52432 to study the modulation of presynaptic GABA_Breceptor on glutamatergic synapses function. (2)Using holographic photostimulation and whole-cell recording to bypass the GABA_Breceptors on presynaptic terminals,selectively research the role of postsynaptic GABA_BRs and glutamate.(3)Using Golgi-staining methods to analyze the morphological classification of SG neurons by sagittal and coronal cutting.Results(1)Under the condition of the sub-threshold stimulus intensity which produced no detectable response,blocking presynaptic GABA_BRs by CGP52432 switched“silent”synapses to successful ones;In the paired-pulse stimulation in which the first stimulation produced about 50%detectable events,CGP52432 increased the amplitude of the evoked excitatory postsynaptic currents(e EPSCs),improving the success rate of e EPSCs,and change the paired-pulse ratio.(2)A presynaptic stimulation train induced a slow excitatory membrane current,the amplitude of the slow current increased in the presence of CGP52432;Tiagabine,a GABA transporter 1 inhibitor,reduced the amplitude of e EPSCs,the addition of CGP52432 restored the amplitude of e EPSCs.(3)Using holographic photostimulation and uncaged glutamate,in the presence of GABA_BRs agonist baclofen(10μm),decrease the amplitude of uncaging excitatory postsynaptic potentials(u EPSPs).Activating different dendrites from the same soma resulted in u EPSPs integration.In the presence of baclofen,the integration significantly decreased.(4)Using Golgi staining,on the sagittal and coronal sections of the spinal cord,it was observed that the volume of neurons in the SG was small,and the boundary with layer III was not obvious.The dendrites of some neurons would extend to the outside or inside beyond the scope of the SG;The density distribution of dendritic spines in different neurons is different.Conclusions(1)Blocking presynaptic GABA_BRs can activate glutamatergic synapses and enhance the release of glutamate while increasing the concentration of environmental GABA can reduce the release of presynaptic glutamate.(2)Postsynaptic GABA_BRs alter glutamate response:regulate glutamate-induced u EPSPs and integrate glutamate-induced u EPSPs from different dendrites.(3)Golgi staining identified the morphological characteristics in the coronal and sagittal sections of the SG neurons,and the differences in the different neuronal morphology further suggest that SG neurons play different roles in transmitting and regulating nociception.