␥-aminobutyric acid type A (GABA A ) receptors comprise a subfamily of ligand-gated ion channels whose activity can be modulated by ligands acting at the benzodiazepine binding site on the receptor. The benzodiazepine binding site was characterized using a site-directed mutagenesis strategy in which amino acids of the ␣ 5 subunit were substituted by their corresponding ␣ 1 residues. Given the high affinity and selectivity of ␣ 1 -containing compared with ␣ 5 -containing GABA A receptors for zolpidem, mutated ␣ 5 subunits were co-expressed with ␤ 2 and ␥ 2 subunits, and the affinity of recombinant receptors for zolpidem was measured. One ␣ 5 mutant (bearing P162T, E200G, and T204S) exhibited properties similar to that of the ␣ 1 subunit, notably high affinity zolpidem binding and potentiation by zolpidem of GABA-induced chloride current. Two of these mutations, ␣ 5 P162T and ␣ 5 E200G, might alter binding pocket conformation, whereas ␣ 5 T204S probably permits formation of a hydrogen bond with a proton acceptor in zolpidem. These three amino acid substitutions also influenced receptor affinity for CL218872. Our data thus suggest that corresponding amino acids of the ␣ 1 subunit, particularly ␣ 1 -Ser 204, are the crucial residues influencing ligand selectivity at the binding pocket of ␣ 1 -containing receptors, and a model of this binding pocket is presented.

As a consequence of impaired glucose or fatty acid metabolism, bioenergetic stress in skeletal muscles may trigger myopathy and rhabdomyolysis. Genetic mutations causing loss of function of the LPIN1 gene frequently lead to severe rhabdomyolysis bouts in children, though the metabolic alterations and possible therapeutic interventions remain elusive. Here, we show that lipin1 deficiency in mouse skeletal muscles is sufficient to trigger myopathy. Strikingly, muscle fibers display strong accumulation of both neutral and phospholipids. The metabolic lipid imbalance can be traced to an altered fatty acid synthesis and fatty acid oxidation, accompanied by a defect in acyl chain elongation and desaturation. As an underlying cause, we reveal a severe sarcoplasmic reticulum (SR) stress, leading to the activation of the lipogenic SREBP1c/SREBP2 factors, the accumulation of the Fgf21 cytokine, and alterations of SR–mitochondria morphology. Importantly, pharmacological treatments with the chaperone TUDCA and the fatty acid oxidation activator bezafibrate improve muscle histology and strength of lipin1 mutants. Our data reveal that SR stress and alterations in SR–mitochondria contacts are contributing factors and potential intervention targets of the myopathy associated with lipin1 deficiency.

No abstract

No abstract