The development of abnormal primary sensory neuron excitability and neuropathic pain symptoms after peripheral nerve injury is associated with altered expression of voltage-gated sodium channels (VGSCs) and a modification of sodium currents. To investigate whether the ␤2 subunit of VGSCs participates in the generation of neuropathic pain, we used the spared nerve injury (SNI) model in rats to examine ␤2 subunit expression in selectively injured (tibial and common peroneal nerves) and uninjured (sural nerve) afferents. Three days after SNI, immunohistochemistry and Western blot analysis reveal an increase in the ␤2 subunit in both the cell body and peripheral axons of injured neurons. The increase persists for Ͼ4 weeks, although ␤2 subunit mRNA measured by real-time reverse transcription-PCR and in situ hybridization remains unchanged. Although injured neurons show the most marked upregulation, ␤2 subunit expression is also increased in neighboring non-injured neurons and a similar pattern of changes appears in the spinal nerve ligation model of neuropathic pain. That increased ␤2 subunit expression in sensory neurons after nerve injury is functionally significant, as demonstrated by our finding that the development of mechanical allodynia-like behavior in the SNI model is attenuated in ␤2 subunit null mutant mice. Through its role in regulating the density of mature VGSC complexes in the plasma membrane and modulating channel gating, the ␤2 subunit may play a key role in the development of ectopic activity in injured and non-injured sensory afferents and, thereby, neuropathic pain.

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1. A method was developed which allows the rapid preparation of pure plasma membranes in high yield from cultured fibroblasts. 2. Cells are lysed in hypo-osmotic borate/EDTA and, after differential centrifugation, the membranes collected by centrifugation on a sucrose barrier. 3. Electron microscopy of the isolated material shows large membrane vesicles essentially free from contaminating organelles. 4. There is no detectable activity of the endoplasmic-reticulum enzyme marker, NADH2—lipoamide oxidoreductase (EC 1.6.4.3), and that of succinate dehydrogenase (EC 1.3.99.1), a marker for mitochondria, is substantially decreased. Chemical compositions are in good agreement with previous observations. 5. This study confirms the usefulness of applied isotopic markers for isolating plasma membranes.