Meeting Abstract
Crassostrea virginica contain serotonin and dopamine, which mediate physiologic functions in gills and other organs. Histamine (HIS), a biogenic amine neurotransmitter in nervous systems and sensory receptors in invertebrates, has rarely been reported in bivalves. Recently, we showed in C. virginica HIS is involved in sensory reception in sensory-motor integration of gill lateral cell cilia beating. We used HPLC to quantify HIS and immunohistofluorescence to detect HIS and HIS H2 receptors in tissues including interfilamental junctions of gill. We hypothesize H2 receptors could be confirmed by Western Blot (WB) analysis, and HIS has a physiological action on interfilamental junctions. For WB gill lysate was prepared by polytron disruption in NP-40 detergent buffer containing protease inhibitor, then centrifuged to obtain supernatant of solubilized membrane proteins. Up to 30 µg of solubilized protein was subjected to SDS-PAGE with 10% acrylamide gels and electroblotted onto nitrocellulose. H2 receptor immunoreactivity was revealed with primary and HRP-conjugated secondary antibodies, and developed using CN/DAB substrate kit. Physiological effects of HIS at interfilamental junctions were observed with a Leica microscope. Responses of interfilamental junctions to HIS and the HIS antagonist famotidine were photographed. WB showed a strong band at 70 kD corresponding to H2 receptors. The physiology study showed HIS (10-3 – 10-5 M) caused a dose-dependent contraction of interfilamental junctions. Famotidine (10-3 – 10-5 M) blocked contractions. The study confirms previous immunohistofluorescence findings of the presence of H2 receptors in gill of C. virginica and identifies a specific physiological role of HIS in the gill.
