ACETYLCHOLINE-CONTAINING NEUROEPITHELIAL CELLS IN FISH GILLS SUPPORT THE CHOLINERGIC HYPOTHESIS OF O2 CHEMORECEPTION
Clinton Wanner, Mark L. BurlesonDepartment of Biological Sciences, College of Arts and Sciences
WHY IS O2 IMPORTANT?
Human survival: 3 weeks without food 3 days without water 3 minutes without oxygen
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200
160
120
80
40
240ox
ygen
par
tial p
ress
ure
(torr)
Environmental O2 Availability:Dissolved Oxygen in Trinity River, Ft. Worth
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9650
5280
2290
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-2000
Everest
Dead Sea
DISCOVERY OF O2 CHEMORECEPTORS Corneille Heymans
Nobel Prize in Physiology or Medicine in 1938 Chemoreceptors in cardio-aortic and carotid
sinus areas Showed that chemical changes in arterial blood
elicited cardio-ventilatory responses. Despite nearly 75 years of research, we still do
not understand how O2 chemoreceptors work!
WHAT ARE O2 CHEMORECEPTORS? O2 chemoreceptors are specialized cells that
detect changing levels of O2 availability and demand and initiate cardiovascular and ventilatory reflexes to maintain normal O2 uptake.
From Lahiri et al., 2001
From Lahiri et al., 2001
WHAT ARE O2 CHEMORECEPTORS?Hypoxia (low oxygen) depolarizes the O2 chemoreceptor causing the release of a neurotransmitter.
The goal of this study is to identify that neurotransmitter using a non-mammalian animal model.
The branchial neuroepithelial cells of fish gills are the evolutionary precursors to mammalian O2 receptors.
Histochemical Profiles of O2 Chemoreceptor Cells
Mammalsserotonindopaminenorepinephrineacetylcholineepinephrinesubstance-Penkephalinsnitric oxideneuron-specific enolasetyrosine hydroxylase
Birdsserotoninsubstance-Pneuron-specific enolasetyrosine hydroxylaseReptilesserotoninenkephalinsneuron-specific enolasetyrosine hydroxylase
Amphibiansserotoninenkephalinsneuron-specific enolasetyrosine hydroxylaseFishserotoninenkephalinsneuron-specific enolasetyrosine hydroxylase
There is no consensus on the roles of any of these chemicalsin the chemoreceptor control of ventilation in vertebrates!
PHYLOGENY Evolution of air-breathing was accompanied
by a reduction and internalization of O2-sensitive chemoreceptors and their loci.
IXX IXX X IX
Fish Amphibians Birdsnon-ChelonianReptiles
Mammals
Fig. 1
HYPOTHESIS : BRANCHIAL NEUROEPITHELIAL CELLS CONTAIN ACETYLCHOLINE.
This hypothesis will be tested using immunohistochemistry and laser confocal microscopy
•Presence of the chemical within the cell. The chemical is either synthesized by the neuron or is taken up from other cells that release it.2
•Stimulus-dependent release. It is released in appropriate quantities by the neuron upon stimulation.
•Action on postsynaptic cell. Exogenous application of the substance in appropriate amounts mimics the action of the endogenously-released substance on the postsynaptic cell or organ.
Mechanism for removal. [Note, not always included as a criterion] A specific mechanism exists to remove the substance from the synaptic cleft, i.e., by degradation or reuptake.
CRITERIA FOR NEUROTRANSMITTER:
PREVIOUS STUDIES: Only acetylcholine consistently mimics the effects
of hypoxia and cyanide (histotoxic hypoxia) on cardio-ventilatory reflex responses and neural activity in mammals and fish.
Furthermore, the effects are mediated by the nicotinic cholinergic receptor subtype.
MATERIAL AND METHODS Channel catfish (Ictalurus punctatus)
obtained from TP&W Dundee Fish Hatchery Maintained in 100 gal tanks. On day of experiment, fish anesthetized
(MS222), heparinzed, cannulated and gills exsanguinated.
Pieces of gill with filaments removed with scissors and fixed in buffered formadehyde
Processed, mounted, stained. Observed using Zeiss 200M inverted optical
microscope modified for confocal laser microscopy.
CONCLUSIONBranchial neuroepithelial of catfish
gills cells contain acetylcholine.
These data support previous reflex and neural studies implicating acetylcholine as the principal neurochemical link between O2-sensitve chemoreceptor cells and primary sensory afferent nerves.