Unexpected association of the “inhibitory” neuroligin 2 with excitatory PSD95 in neuropathic pain

Abstract

In the spinal nerve ligation (SNL) model of neuropathic pain, synaptic plasticity shifts the excitation/inhibition balance toward excitation in the spinal dorsal horn. We investigated the deregulation of the synaptogenic neuroligin (NL) molecules, whose NL1 and NL2 isoforms are primarily encountered at excitatory and inhibitory synapses, respectively. In the dorsal horn of SNL rats, NL2 was overexpressed whereas NL1 remained unchanged. In control animals, intrathecal injections of small interfering RNA (siRNA) targeting NL2 increased mechanical sensitivity, which confirmed the association of NL2 with inhibition. By contrast, siRNA application produced antinociceptive effects in SNL rats. Regarding NL partners, expression of the excitatory postsynaptic scaffolding protein PSD95 unexpectedly covaried with NL2 overexpression, and NL2/PSD95 protein interaction and colocalization increased. Expression of the inhibitory scaffolding protein gephyrin remained unchanged, indicating a partial change in NL2 postsynaptic partners in SNL rats. This phenomenon appears to be specific to the NL2(-) isoform. Our data showed unexpected upregulation and pronociceptive effects of the "inhibitory" NL2 in neuropathic pain, suggesting a functional shift of NL2 from inhibition to excitation that changed the synaptic ratio toward higher excitation.

Neuroligin 2 regulates spinal GABAergic plasticity in hyperalgesic priming, a model of the transition from acute to chronic pain

“…Moreover, these investigators also found that nlgn2 knockdown, in their case with shRNA, led to an alleviation of mechanical hypersensitivity brought about by peripheral nerve injury yet caused mechanical hypersensitivity in sham rats. 15 This is strikingly similar to our findings with the nlgn2 interfering peptide, neurolide 2. Interestingly, while the study in rats with peripheral nerve injury found an upregulation of the nlgn2(−) splice variant in the spinal cord, 15 we observed an increase in nlgn2A.…”

“…10 Differential expression of these splice isoforms have also been implicated in neuropathic pain. 15 Therefore, we assessed nlgn2 mRNA levels, including these splice variants, using qPCR on spinal dorsal horn and DRG total RNA extracts. We administered i.t.…”

“…We tested the effect of neurolide 2 on naive mice, where we predict that inhibition of nlgn2 function should induce mechanical hypersensitivity, consistent with observation in rats using shRNA. 15 Neurolide 2 i.t. injection caused robust mechanical hypersensitivity in naive mice that resolved within 7 days (Fig.…”

“…A very recent study suggested a novel mechanism of dorsal horn plasticity, alterations in nlgn expression and function that may play an important role in neuropathic pain. 15 This led us to explore the possible role of nlgn1 and 2 plasticity in the setting of hyperalgesic priming. Our findings support a model wherein nlgn2 controls pathological alterations in inhibitory synapse structure and function that underlies the maintenance of hyperalgesic priming.…”

“…1 An underlying feature of these changes may be differences in postsynaptic plasticity between inhibitory and excitatory neurons in response to afferent input, 27 decreased expression or function of the neuron-specific K + , Cl − cotransporter 2 (KCC2), 14,23,38,39 and/or altered expression of the inhibitory synapse adhesion molecule neuroligin 2 (nlgn2). 15 KCC2 establishes low intracellular Cl − concentrations that are required for inhibitory hyperpolarization on opening of GABA A Rs. Loss of KCC2 expression after injury is associated with the development of mechanical allodynia and the generation of neuropathic pain.…”