PT - JOURNAL ARTICLE AU - Hidehiko Koizumi AU - Bryan Mosher AU - Mohammad F. Tariq AU - Ruli Zhang AU - Naohiro Koshiya AU - Jeffrey C. Smith TI - Voltage-dependent rhythmogenic property of respiratory pre-Bötzinger complex glutamatergic, Dbx1-derived and somatostatin-expressing neuron populations revealed by graded optogenetic inhibition AID - 10.1523/ENEURO.0081-16.2016 DP - 2016 May 25 TA - eneuro PG - ENEURO.0081-16.2016 4099 - http://www.eneuro.org/content/early/2016/05/25/ENEURO.0081-16.2016.short 4100 - http://www.eneuro.org/content/early/2016/05/25/ENEURO.0081-16.2016.full AB - The rhythm of breathing in mammals, originating within the brainstem pre-Bötzinger complex (pre-BötC), is presumed to be generated by glutamatergic neurons, but this has not been directly demonstrated. Additionally, developmental expression of the transcription factor, Dbx1, or expression of the neuropeptide somatostatin (Sst), has been proposed as a marker for the rhythmogenic pre-BötC glutamatergic neurons, but it is unknown if these other two phenotypically-defined neuronal populations are functionally equivalent to glutamatergic neurons with regard to rhythm generation. To address these problems, we comparatively investigated, by optogenetic approaches, the roles of pre-BötC glutamatergic, Dbx1-derived, and Sst-expressing neurons in respiratory rhythm generation in neonatal transgenic mouse medullary slices in vitro and also more intact adult perfused brainstem-spinal cord preparations in situ. We established three different triple transgenic mouse lines with Cre-driven Archaerhodopsin-3 (Arch) expression selectively in glutamatergic, Dbx1-derived, or Sst-expressing neurons for targeted photoinhibition. In each line, we identified subpopulations of rhythmically active, Arch-expressing pre-BötC inspiratory neurons by whole-cell recordings in medullary slice preparations in vitro, and established that Arch-mediated hyperpolarization of these inspiratory neurons was laser power-dependent with equal efficacy. By site- and population-specific graded photoinhibition, we then demonstrated that inspiratory frequency was reduced by each population with the same neuronal voltage-dependent frequency control mechanism in each state of the respiratory network examined. We infer that enough of the rhythmogenic pre-BötC glutamatergic neurons also have the Dbx1- and Sst-expression phenotypes and thus all three phenotypes share the same voltage-dependent frequency control property.Significance Statement: The brainstem pre-Bötzinger complex (pre-BötC) is the origin of rhythmic neural activity producing inspiratory movements in mammals. Despite over two decades of investigation, there is the central problem of establishing the causal role of specific pre-BötC neuron populations in generating this rhythmic motor behavior. By optogenetic and electrophysiological approaches, we reveal that excitatory glutamatergic, transcription factor Dbx1-derived, and somatostatin-expressing neuronal populations, overlapping within the pre-BötC region, are functionally equivalent in respiratory rhythm generation with the same neuronal voltage-dependent frequency control mechanism in different functional and developmental states of the respiratory network including neonatal in vitro and adult in situ conditions. Our results help resolve the longstanding problem of delineating the relative roles of these excitatory interneuron populations in rhythm generation.