The gaseous messenger molecule, nitric oxide : a modulator of locomotor movements during early amphibian development
Abstract
1. The free radical gas nitric oxide (NO) is now recognised as a ubiquitous and versatile
signalling molecule and the investigation of its biological roles has involved a wide
range of scientific disciplines in many different species. Yet despite this, its potential
roles in the development of rhythmic motor activities in vertebrates have been largely
ignored.
2. Physiological experiments recording extracellular ventral root output suggest that NO is
playing an inhibitory role in the swimming system of Xenopus laevis larvae, shortening
the duration of swim episodes and slowing swim frequency. Nicotinamide adenine
dinucleotide phosphate (NADPH)-diaphorase histochemistry labelled three populations
of neurons in the brainstem, which putatively co-localise NO with the aminergic
neuromodulators serotonin (5-HT) and noradranaline (NA), and the fast descending
inhibitory neurotransmitter, y-aminobutyric acid (GABA). This suggests that the
inhibitory role is supraspinal in origin.
3. Intracellular recordings from neurons presumed to be spinal motor neurons provide
further evidence for the inhibitory influence of NO. My experiments suggest that NO
potentiates both glycinergic and y-aminobutyric acid (GABA)-ergic inhibition onto
spinal motor neurons. The facilitation of the release of these inhibitory transmitters is
consistent with the observed effects on swim frequency and swim episode duration,
respectively. Additionally, NO appears to affect membrane properties, causing a
pronounced membrane potential depolarisation and a decrease in membrane
conductance. This suggests that NO shuts off a resting membrane conductance.
4. NADPH-diaphorase histochemistry was subsequently applied to determine the four
dimensional expression of putative nitrergic neurons in the central nervous system and
related structures. The developmental sequence of staining identifies groups and
subgroups of interconnected intemeurons, and provides further clues to their identity.
NADPH-diaphorase labelling was also located in the eyes, skin and blood vessels,
further confirming the validity of this staining technique for identirying nitric oxide
synthase.
5. In the related anuran species, Rana temporaria nitric oxide donor drugs appear to have
no affect on swimming, but instead reliably initiates a non-rhythmic "lashing" motor
pattern similar to that elicited by dimming of the illumination. Interestingly the
NADPH-diaphorase technique labelled three clusters of apparently homologous
interneurons in the brainstem and additionally the inner layer of the skin was intensely
stained, implicating a species-specific role for NO released from brainstem neurons.
Type
Thesis, PhD Doctor of Philosophy
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