The Bovine Ex Vivo Retina: A Versatile Model for Retinal Neuroscience.

Purpose: The isolated ex vivo retina is the standard model in retinal physiology and neuroscience. During isolation, the retina is peeled from the retinal pigment epithelium (RPE), which plays a key role in the visual cycle. Here we introduce the choroid-attached bovine retina as an in vivo-like model for retinal physiology. We find that-in the bovine eye-the choroid and retina can be peeled from the sclera as a single thin sheet. Importantly, the retina remains tightly associated with the RPE, which is sandwiched between the retina and the choroid. Furthermore, bovine tissue is readily available and cheap, and there are no ethical concerns related to the use of animals solely for research purposes. Methods: We combine multi-electrode array and single-cell patch-clamp recordings to characterize light responses in the choroid-attached bovine ex vivo retina. Results: We demonstrate robust and consistent light responses in choroid-attached preparations. Importantly, light responses adapt to different levels of background illumination and rapidly recover from photobleaching. The choroid-attached retina is also thin enough to permit targeted electrophysiological recording from individual retinal neurons using standard differential interference contrast microscopy. We also characterize light responses and membrane properties of bovine retinal ganglion cells and compare data obtained from bovine and murine retinas. Conclusions: The choroid-attached retinal model retains the advantages of the isolated retina but with an intact visual cycle and represents a useful tool to elucidate retinal physiology.

Modulation of GABAergic Synaptic Transmission by NMDA Receptors in the Dorsal Horn of the Spinal Cord.

The dorsal horn (DH) of the spinal cord is an important structure involved in the integration of nociceptive messages. Plastic changes in the properties of neuronal networks in the DH underlie the development of analgesia as well as of hyperalgesia and allodynia in acute and chronic pain states. Two key mechanisms are involved in these chronic pain states: increased electrical activities and glutamate release leading to the recruitment of NMDAr and plastic changes in the synaptic inhibition. Although: (1) the balance between excitation and inhibition is known to play a critical role in the spinal network; and (2) plastic changes in spinal excitation and inhibition have been studied separately, the relationship between these two mechanisms has not been investigated in detail. In the present work, we addressed the role of NMDA receptors in the modulation of GABAergic synaptic transmission in the DH network. Using tight-seal whole-cell recordings on adult mice DH neurons, we characterized the effect of NMDAr activation on inhibitory synaptic transmission and more especially on the GABAergic one. Our results show that, in a subset of neurons recorded in lamina II, NMDAr activation facilitates spontaneous and miniature GABAergic synaptic transmission with a target specificity on GABAergic interneurons. In contrast, NMDA reduced the mean amplitude of evoked GABAergic IPSCs. These results show that NMDAr modulate GABAergic transmission by a presynaptic mechanism of action. Using a pharmacological approach, we investigated the composition of NMDAr involved in this modulation of GABAergic synaptic transmission. We found that the NMDA-induced facilitation was mediated by the activation of NMDAr containing GluN2C/D subunits. Altogether, our results bring new insights on nociceptive information processing in the spinal cord network and plastic changes in synaptic inhibition that could underlie the development and maintenance of chronic pain.

Inhibitory interneurons with differential plasticities at their connections tune excitatory-inhibitory balance in the spinal nociceptive system.

ABSTRACT: Networks of the dorsal horn of the spinal cord process nociceptive information from the periphery. In these networks, the excitation-inhibition balance is critical to shape this nociceptive information and to gate it to the brain where it is interpreted as pain. Our aim was to define whether short-term plasticity of inhibitory connections could tune this inhibition-excitation balance by differentially controlling excitatory and inhibitory microcircuits. To this end, we used spinal cord slices from adult mice expressing enhanced green fluorescent protein (eGFP) under the GAD65 promoter and recorded from both eGFP+ (putative inhibitory) and eGFP- (putative excitatory) neurons of lamina II while stimulating single presynaptic GABAergic interneurons at various frequencies. Our results indicate that GABAergic neurons of lamina II simultaneously contact eGFP- and eGFP+ neurons, but these connections display very different frequency-dependent short-term plasticities. Connections onto eGFP- interneurons displayed limited frequency-dependent changes and strong time-dependent summation of inhibitory synaptic currents that was however subjected to a tonic activity-dependent inhibition involving A1 adenosine receptors. By contrast, GABAergic connections onto eGFP+ interneurons expressed pronounced frequency-dependent depression, thus favoring disinhibition at these synapses by a mechanism involving the activation of GABAB autoreceptors at low frequency. Interestingly, the balance favors inhibition at frequencies associated with intense pain, whereas it favors excitation at frequencies associated with low pain. Therefore, these target-specific and frequency-specific plasticities allow to tune the balance between inhibition and disinhibition while processing frequency-coded information from primary afferents. These short-term plasticities and their modulation by A1 and GABAB receptors might represent an interesting target in pain-alleviating strategies.