Developmental and regional expression of NADPH-diaphorase/nitric oxide synthase in spinal cord neurons correlates with the emergence of limb motor networks in metamorphosing Xenopus laevis.

Sankari Ramanathan, Denis Combes, Micol Molinari, John Simmers, Keith T. Sillar
European Journal of Neuroscience. 2006-10-01; 24(7): 1907-1922
DOI: 10.1111/j.1460-9568.2006.05057.x

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Ramanathan S(1), Combes D, Molinari M, Simmers J, Sillar KT.

Author information:
(1)School of Biology, University of St Andrews, Bute Medical Buildings, St
Andrews, Fife, United Kingdom, KY16 9TS.

Metamorphosis in anuran amphibians requires a complete transformation in
locomotor strategy from undulatory tadpole swimming to adult quadrupedal
propulsion. The underlying reconfiguration of spinal networks may be influenced
by various neuromodulators including nitric oxide, which is known to play an
important role in CNS development and plasticity in diverse species, including
metamorphosis of amphibians. Using NADPH-diaphorase (NADPH-d) staining and
neuronal nitric oxide synthase (nNOS) immunofluorescence labelling, the
expression and developmental distribution of NOS-containing neurons in the
spinal cord and brainstem were analysed in all metamorphic stages of Xenopus
laevis. Wholemount preparations of the spinal cord from early stages of
metamorphosis (coincident with emergence of the fore- and hindlimb buds)
revealed two clusters of NOS-positive neurons interspersed with areas devoid of
stained somata. These cells were distributed in three topographic subgroups, the
most ventral of which had axonal projections that crossed the ventral
commissure. Motoneurons innervating the fore- and hindlimb buds were
retrogradely labelled with horseradish peroxidase (HRP) to determine their
position in relation to the two NOS-expressing cord regions. Limb motoneurons
and NOS-positive cells did not overlap, indicating that during early stages of
metamorphosis nitrergic neurons are excluded from regions where spinal limb
circuits are forming. As metamorphosis progresses, NOS expression became
distributed along the length of the spinal cord together with an increase in the
number and intensity of labelled cells and fibers. NOS expression reached a peak
as the forelimbs emerge then declined. These findings are consistent with a role
for nitric oxide (NO) in the developmental transition from undulatory swimming
to quadrupedal locomotion.

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