nNOS regulates ciliated cell polarity, ciliary beat frequency, and directional flow in mouse trachea.

Clearance of the airway is dependent on directional mucus flow across the mucociliary epithelium, and deficient flow is implicated in a range of human disorders. Efficient flow relies on proper polarization of the multiciliated cells and sufficient ciliary beat frequency. We show that NO, produced by nNOS in the multiciliated cells of the mouse trachea, controls both the planar polarity and the ciliary beat frequency and is thereby necessary for the generation of the robust flow. The effect of nNOS on the polarity of ciliated cells relies on its interactions with the apical networks of actin and microtubules and involves RhoA activation. The action of nNOS on the beat frequency is mediated by guanylate cyclase; both NO donors and cGMP can augment fluid flow in the trachea and rescue the deficient flow in nNOS mutants. Our results link insufficient availability of NO in ciliated cells to defects in flow and ciliary activity and may thereby explain the low levels of exhaled NO in ciliopathies

Behavior of adolescents of different experimental groups in the social interactions test.

<p>SDS – social defeat stress; PS – psychological stress; Light columns - empty cylinder; dark columns -unfamiliar partner is under cylinder. ***P<0.001 <i>vs</i> the controls of respective partners; +++P<0.001 <i>vs</i> SDS adolescents of respective partners; ##P<0.01; ###P<0.001 <i>vs</i> empty cylinder in respective groups (n = 10–12 per group).</p

Behavior of adolescents of different experimental groups in the partition test.

<p>SDS – social defeat stress; PS – psychological stress; Light columns - familiar partner; dark columns - unfamiliar partner; *P<0.05; **P<0.01; ***P<0.001 <i>vs</i> the controls of respective partners; +P<0.05; ++P<0.01; +++P<0.001 <i>vs</i> SDS adolescents of respective partners; #P<0.05; ##P<0.01; ###P<0.001 <i>vs</i> familiar partner in respective group (n = 10–12 per group).</p

Protocols for studying the effects of combined stress (SDS+SIS or CDS+SIS) on the behavior of adolescents (A) and adult mice stressed in adolescence (B).

<p>SDS - social defeat stress; CDS – communication deprivation stress; SIS – social instability stress. As a control age-matched groups of males living in littermate groups were used. Before being examined in a range of behavioral tests, control animals and adult mice at the period of 31–38 days (B) were placed in individual cages to facilitate behavioral testing and remove group housing effects. Experimental groups and age-matched control groups were tested simultaneously in the behavioral tests (one test per day). Details of protocol are described in section “Materials and methods”.</p

BrdU-positive cells in hippocampal subgranular zone of the dentate gyrus in SDS adolescents and in grown adolescents after 3 weeks of the rest.

<p>SDS – social defeat stress **P<0.01 - <i>vs</i> the controls (n = 6–7 per group).</p

Behavior of adolescents in the elevated plus-maze test.

<p>*P<0.05; **P<0.01 <i>vs</i> the control; ++P<0.01 <i>vs</i> SDS adolescents (n = 10–12 per group).</p

Behavior of adolescents of different experimental groups in the Porsolt test.

<p>SDS – social defeat stress; PS – psychological stress; **P<0.01 <i>vs</i> the controls. #P<0.05 <i>vs</i> SDS adolescents (n = 10–12 per group).</p

Behavior of SDS adolescents in behavioral tests after 3 weeks of rest.

<p>*P<0.05; **P<0.01; ***P<0.001 <i>vs</i> the control; ^#P<0.01; ^##P<0.001 <i>vs</i> empty cylinder.</p

Behavior of adolescents (the controls and group of adolescents after 3 weeks of rest in the elevated plus-maze test.

<p>*P<0.05; **P<0.01; ***P<0.001 <i>vs</i> the controls (n = 11–12 per group).</p