Beating of Gill Lateral Cell Cilia of Crassostrea virginica Involves Neuronal Innervation and the Presence of Gap Junctions
Authors: Buchanan Reniece , Dobey Nicole , Frank Dane , Catapane Edward J , Carroll Margaret A 
Published: November 2023

Suspension feeding bivalves use a ciliary pump driven by tracts of lateral cilia located on gill filaments to provide water currents for feeding, respiration and waste elimination. These cells in Crassostrea virginica are controlled by serotonergic‐dopaminergic innervation via the branchial nerve. Serotonin is the neurotransmitter that increases beating rates, while dopamine decreases it. Lateral cells of many other bivalves are similarly innervated. The cellular mechanism regulating lateral cell cilia beating is not fully explained. Second messenger systems are involved and gill filaments receive branches from the branchial nerve, but it is not known if each lateral cell is individually innervated. Lateral cells have gap junctions and questions arise if gap junctions are involved in controlling the rate or coordination of beating. We hypothesize gap junctions are involved in the cellular mechanism regulating lateral cell cilia beating rate and/or coordination to maintain the metachronal wave pattern. To study this we made immunohistological preparations using antibodies to visualize the gap junctions, and used the Sihler Whole Mount method to visualize nerves in gill filaments. We also set up preparations in which gill filaments were divided into proximal and distal sections by a petroleum jelly barrier, which prevented diffusion of drugs from one side to the other. We stimulated the branchial nerve entering the proximal end of the filaments with suction electrodes (5 hz, 2 msec, 10 – 20 volts), which causes a release of serotonin and increases beating rates. Stroboscopic microscopy was used to measure cilia beating rates. Drugs were applied to distal ends of preparations and beating rates of proximal and distal ends were compared. Applying procaine, a local anesthetic, to distal ends prevented electrical stimulation from increasing beating rates, while beating rate at the proximal ends increased, showing nerve impulse propagation is necessary for ciliary response. Using the same procedure, we tested 3 gap junction blockers, each with different mechanisms of action: lindane, diphenylborinic anhydride and mefloquine. Mefloquine had no effect on beating, but lindane and diphenylborinic anhydride both produced statistically significant reductions in beating rates at the distal ends. Lindane was stronger, causing a dose‐dependent (10−7 – 10−4M) blockage of beating, while diphenylborinic anhydride only blocked at a high dose of 10−4M. Results show neuronal innervation to filaments in conjunction with functioning gap junctions in the cells are necessary to increase cilia beating rates. Disrupting either one interfered with the normal response. Lindane is reported to reduce gap junction permeability and increase intracellular Ca2+ levels; either one or both of these actions may be responsible for the results we obtained.Support or Funding InformationThis work was supported in part by 2R25GM06003 of the Bridge Program of NIGMS and 0516041071 of NYSDOE.