ASH activity is associated with locomotion arousal.
2015-07-08T02:55:35Z (GMT) by
<p>Locomotion behavior during the L4/A lethargus (A-C) and in adults (D) of single worms whose ASH neurons were ablated by transgenic overexpression of CED-3 in ASH neurons (<i>sra-6</i> promoter) was analyzed in the indicated genotypes. Animals were analyzed by fluorescence microscopy after locomotion recordings to determine if ASH neurons were ablated (1–2 ASH: animals with 1 or 2 ASH intact neurons; 0 ASH: animals lacking viable ASH neurons). Instantaneous locomotion velocity (A), average motile fraction (B), and average locomotion velocity (C-D) are plotted. The <i>npr-1</i> locomotion defect during the L4/A lethargus, but not in adults, was partially suppressed in the transgenic animals in which both of ASH neurons were ablated (0 ASH). (E-H) Copper-evoked calcium transients in ASH were analyzed in L4, L4/A, and adults of the indicated genotypes using cameleon as a calcium indicator. Averaged responses (E, G), and the amplitudes of individual trials (F, H) are shown for each genotype. Each trace represents the average percentage change in YFP/CFP fluorescence ratio. The light tan rectangle indicates the duration for which 10 mM copper was applied. Dark gray shading of each trace indicates SEM of the mean response. (E-F) Copper-evoked calcium transients in ASH neurons were significantly reduced during L4/A lethargus, and this effect was abolished in <i>npr-1</i> mutants. (G-H) This defect during L4/A lethargus was rescued by transgenes expressing NPR-1 in the RMG circuit (RMG rescue, <i>flp-21 promoter</i>) or in ASH neurons (ASH rescue, <i>sra-6 promoter</i>). (I-J) Forced depolarization of ASH neurons increased adult locomotion velocity (I) and aldicarb sensitivity (J). Rat TRPV1 was ectopically expressed in ASH neurons (using the <i>sra-6</i> promoter). (I) Locomotion behavior of adult transgenic worms was analyzed with or without capsaicin treatment (5 hours). Average locomotion velocity (I) is plotted. Capsaicin treatment increased adult locomotion velocity in transgenic animals expressing TRPV1 in ASH neurons, but not in wild type controls. The number of animals analyzed is indicated for each genotype. (J) The percentage of animals paralyzed on 1 mM aldicarb at 80 min with or without capsaicin treatment (2–3 hours pretreatment) were plotted for the indicated genotypes. The number of trials is indicated for each genotype. Full time courses for aldicarb-induced paralysis are shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005359#pgen.1005359.s002" target="_blank">S2G Fig</a>. Capsaicin treatment increased aldicarb sensitivity in transgenic animals expressing TRPV1 in ASH neurons, but not in wild type controls. Error bars indicate SEM. Values that differ significantly are indicated (***, <i>p</i> <0.001; ns, not significant).</p>