Selective regulation of 3α-hydroxysteroid oxido-reductase expression in dorsal root ganglion neurons: A possible mechanism to cope with peripheral nerve injury-induced chronic pain

The enzyme 3α-hydroxysteroid oxido-reductase (3α-HSOR) catalyzes the synthesis and bioavailability of 3α,5α-neurosteroids as allopregnanolone (3α,5α-THP) which activates GABAA receptors and blocks T-type calcium channels involved in pain mechanisms. Here, we used a multidisciplinary approach to demonstrate that 3α-HSOR is a cellular target the modulation of which in dorsal root ganglia (DRG) may contribute to suppress pain resulting from peripheral nerve injury. Immunohistochemistry and confocal microscope analyses showed 3α-HSOR-immunostaining in naive rat DRG sensory neurons and glial cells. Pulse-chase, high performance liquid chromatography and Flo/One characterization of neurosteroids demonstrated 3α,5α-THP production in DRG. Behavioral methods allowed identification of pain symptoms (thermal and mechanical hyperalgesia and/or allodynia) in rats subjected to sciatic nerve chronic constriction injury (CCI). Reverse transcription and real-time polymerase chain reaction revealed that 3α-HSOR mRNA concentration in CCI-rat ipsilateral DRG, 5-fold higher than in contralateral DRG, was also 4- to 6-fold elevated than that in sham-operated or naive rat DRG. Consistently, Western blotting confirmed increased 3α-HSOR protein levels in CCI-rat ipsilateral DRG and double immunolabeling showed that 3α-HSOR overexpression occurred in DRG neurons but not in glia. Functional plasticity of 3α-HSOR leading to increased 3α,5α-THP production was evidenced in CCI-rat DRG. Interestingly, behavioral and molecular time-course investigations revealed that 3α-HSOR gene upregulation was correlated to pain symptom development. Most importantly, in vivo knockdown of 3α-HSOR expression in healthy rat DRG using 6-carboxyfluorescein-3α-HSOR-siRNA exacerbated thermal and mechanical pain perceptions. This paper is the first to show that siRNA-induced knockdown of a key neurosteroid-synthesizing enzyme directly affects an important function as nociception. Hopefully, these results may be useful for the development of novel analgesics.