Lack of evidence for synaptic high‐affinity γ‐hydroxybutyric acid (GHB) transport in rat brain synaptosomes and 11 Na⁺‐dependent SLC neurotransmitter transporters

Abstract

γ‐Hydroxybutyric acid (GHB) is an endogenous compound proposed to act as a neurotransmitter. Na+‐dependent, high‐affinity GHB transport has long been considered important evidence supporting this hypothesis. However, the molecular identity of such a high‐affinity transporter remains unknown. In this study, we sought to identify and characterize GHB synaptic transport through a series of studies using both native and recombinant systems with the ultimate aim of providing evidence to clarify the proposed role of GHB as a neurotransmitter in the mammalian brain. Native [3H]GHB transport was studied in isolated rat brain synaptosomes and compared to synaptic membranes. As a targeted approach, GHB was also screened against a panel of Na+‐dependent SLC6 neurotransmitter transporters recombinantly expressed in Xenopus laevis oocytes or tsA201 cells. Finally, the low‐affinity GHB transporters, MCT1/2 and SMCT1, were probed as GHB transporters in L‐[14C]lactate uptake assays in synaptosomes. We found no evidence of high‐affinity [3H]GHB transport in purified rat brain cortical or striatal synaptosomes or at any of the 11 SLC6 transporters tested. Instead, our results indicate the binding of [3H]GHB to an unidentified membrane component, distinct from any of the known GHB targets. In accordance with others, we found that GHB and the analog 3‐hydroxycyclopent‐1‐enecarboxylic acid (HOCPCA) can, in millimolar concentrations, inhibit L‐[14C]lactate uptake at MCT1 and/or MCT2 and that this also can occur in synaptosomes. In conclusion, through a variety of in vitro pharmacological studies, we were unsuccessful in identifying a specific synaptic high‐affinity transporter for GHB. Our findings emphasize the need to reevaluate GHB's role as a potential neurotransmitter. Open Science Badges This article has received a badge for *Open Materials* because it provided all relevant information to reproduce the study in the manuscript. The complete Open Science Disclosure form for this article can be found at the end of the article. More information about the Open Practices badges can be found at https://cos.io/our-services/open-science-badges/.