The heterotrimeric G-protein α subunit, Gαolf, acts to transduce extracellular signals through G-protein coupled receptors (GPCRs) and stimulates adenylyl cyclase mediated production of the second messenger cyclic adenosine monophosphate. Numerous mutations in the GNAL gene, which encodes Gαolf, have been identified as causative for an adult-onset dystonia. These mutations disrupt GPCR signaling cascades in in vitro assays through several mechanisms, and this disrupted signaling is hypothesized to lead to dystonic motor symptoms in patients. However, the cells and circuits that mutations in GNAL corrupt are not well understood. Published patterns of Gαolf expression outside the context of the striatum are sparse, conflicting, often lack cell type specificity, and may be confounded by expression of the close GNAL homolog of GNAS. Here, we use RNAScope in-situ hybridization to quantitatively characterize Gnal mRNA expression in brain tissue from wildtype C57BL/6J adult mice. We observed widespread expression of Gnal puncta throughout the brain, suggesting Gαolf is expressed in more brain structures and neuron types than previously accounted for. We quantify transcripts at a single cell level, and use neuron type specific markers to further classify and understand patterns of GNAL expression. Our data suggests that brain regions classically associated with motor control, initiation, and regulation show the highest expression of GNAL, with Purkinje Cells of the cerebellum showing the highest expression of any neuron type examined. Subsequent conditional Gnal knockout in Purkinje cells led to markedly decreased intracellular cAMP levels and downstream cAMP-dependent enzyme activation. Our work provides a detailed characterization of Gnal expression throughout the brain and the biochemical consequences of loss of Gαolf signaling in vivo in neurons that highly express Gnal.

Zinc is recognized as an important element for olfaction. Zinc nanoparticles enhance olfaction in response to odors; however, the mechanisms underlying this action remain unknown. Herein, the effect of zinc on olfactory receptors was deduced using electro-olfactogram (EOG) responses recorded from the isolated olfactory mucosae of bullfrogs (Rana catesbeiana) following the administration or chelation of zinc ions. Menthone and n-amyl acetate were used as odorants, whereas forskolin (an adenylate cyclase activator) and cholera toxin (a Gαolf activator) were used as intracellular signal transduction activators. The EOG responses provoked by the odorants and cholera toxin were suppressed by dithizone-mediated zinc ion chelation, and the EOG responses were recovered by administering non-chelated zinc. However, the EOG response to forskolin was not suppressed by dithizone. In contrast, the addition of femtomolar concentrations of zinc ions enhanced the EOG responses. The above-mentioned effects on EOG responses were examined by changing the concentration of zinc ions but not zinc nanoparticles. The results of this study suggest that Gαolf alone or both olfactory receptors and Gαolf likely require zinc ions for their activation.

GNAL was identified as a cause of dystonia in patients from North America, Europe and Asia. In this study, we aimed to investigate the prevalence of GNAL variants in Brazilian patients with dystonia. Ninety-one patients with isolated idiopathic dystonia, negative for THAP1 and TOR1A mutations, were screened for GNAL variants by Sanger sequencing. Functional characterization of the Gαolf protein variant was performed using the bioluminescence resonance energy transfer assay. A novel heterozygous nonsynonymous variant (p. F133L) was identified in a patient with cervical and laryngeal dystonia since the third decade of life, with no family history. This variant was not identified in healthy Brazilian controls and was not described in 63,000 exomas of the ExAC database. The F133L mutant exhibited significantly elevated levels of basal BRET and severely diminished amplitude of response elicited by dopamine, that both indicate substantial functional impairment of Gαolf in transducing receptor signals, which could be involved in dystonia pathophysiology. GNAL mutations are not a common cause of dystonia in the Brazilian population and have a lower prevalence than THAP1 and TOR1A mutations. We present a novel variant that results in partial Gαolf loss of function.