γ‐Hydroxybutyrate (GHB) is an endogenous neuromodulator with therapeutical applications in anesthesia, sleep disorders, and drug addiction. We report the cloning of a GHB receptor from a rat hippocampal cDNA library. This receptor has a molecular mass of 56 kDa and belongs to the seven‐transmembrane receptor family. The peptidic sequence has no significant homology with any known receptor, including GABA _B receptors. Its mRNA is restricted to the brain and is particularly abundant in the hippocampus, cortex, striatum, thalamus, olfactory bulbs, and cerebellum, matching the distribution of GHB binding sites in rat brain. Southern blot revealed the presence of homologous sequences in several species including the human. Binding assays on transfected CHO cells showed a dissociation constant ( K _d ) of 426 nM for GHB and no affinity for GABA, baclofen, or glutamate. In patch‐clamp experiments, transfected CHO cells revealed a functional G‐protein‐coupled receptor as demonstrated by GTP‐γ‐S‐induced irreversible activation. Application of 0.1‐15 µM GHB specifically induced an inward current at negative membrane potentials that was not reproduced by application of baclofen (10 µM). CGP‐55845, a GABA _B receptor antagonist, did not inhibit the GHB‐induced response nor did the GHB receptor antagonist NCS‐382, suggesting that the GHB receptor system includes several subtypes.

Oxaliplatin (OXAL) is a platinum-based drug used for the treatment of colorectal, lung, breast and ovarian cancers. OXAL does not cause renal or hematologic toxicity. However, OXAL induces neuropathic pain which hampers the chemotherapy success. Attempts with neuroprotective agents including anticonvulsivants and antidepressants were made to prevent OXAL-induced painful neuropathy but the clinical data are controversial and the tested neuroprotectors are able to evoke themselves undesirable effects. Here, we demonstrated that the natural neurosteroid allopregnanolone (3α,5α-THP), known to be devoid of toxic side-effects in humans and experimental models, prevented and suppressed OXAL-induced painful neuropathic symptoms. Indeed, 3α,5α-THP repaired OXAL-evoked neurochemical and functional alterations in peripheral nerves and intra-epidermal nerve fibers (IENF). Behavioral analyses showed that prophylactic or corrective 3α,5α-THP treatment (4mg/kg/2days) respectively prevented or abolished OXAL-induced cold allodynia, mechanical allodynia and hyperalgesia by reversing to normal decreased thermal and mechanical pain thresholds of OXAL-treated rats. Electrophysiological investigations revealed that 3α,5α-THP restored control values of sciatic nerve conduction velocity and action potential peak amplitude drastically reduced by OXAL-treatment. Furthermore, immunohistochemistry and confocal microscopic quantifications demonstrated that 3α,5α-THP repaired OXAL-induced neurochemical/cellular alterations by restoring IENF control density and normal level of neurofilament 200kDa that was strongly repressed by OXAL in dorsal root ganglion neurons and sciatic nerve axons. OXAL showed no toxicity for the non-compact myelin protein 2',3'-cyclic-nucleotide-3'-phosphodiesterase whose expression level was similarly increased by 3α,5α-THP in controls and OXAL-treated rat nerves. Together, these results may be interesting for the development of natural or safe neurosteroid-based neuroprotective strategy against anticancer drug-evoked painful neuropathy.

Two parent clones of a γ‐hydroxybu‐tyrate (GHB) receptor, C12K32 and GHBhl, were isolated from a human frontal cortex cDNA library. The two clones differ by a deleted cytosine in C12K32. CHO cells transfected with either C12K32 or GHBhl responded positively to submicromolar GHB stimulation. However, unlike C12K32, GHBh1 desensitizes rapidly on application of low concentrations of GHB. GHB receptor properties were then studied on C12K32 expressed in CHO cells. C12K32 bound GHB with a K _d of 114 nM and has no affinity for GABA or glutamate. GHB and NCS‐382 displaced [ ³ H]GHB with an IC ₅₀ of 53 ± 8 and 120 ± 18 nM, respectively. In patch‐clamp experiments, GHB induced a dose‐dependent response with an EC ₅₀ of 130 nM. This response was antagonized by NCS‐382, was not reproduced by GABA, and was sensitive to the addition of GTP‐γ‐S in the recording pipette. CHO cells transfected with C12K32 exhibited GTPγ‐ ³⁵ S binding with an EC ₅₀ of 462 nM for GHB and an IC ₅₀ of 2.9 μM for NCS‐382. The present data led to the conclusion that both C12K32 and GHBh1 are two closely related isoforms of a human GHB receptor, GHBh1, that is described in the databank as the GPCR 172A—Andriamampandry, C., Taleb, O., Kemmel, V., Humbert, J.‐P., Aunis, D., Maitre, M. Cloning and functional characterization of a gamma‐hydroxybutyrate receptor identified in the human brain. FASEB J. 21, 885–895 (2007)