Meeting Abstract
P1.145 Sunday, Jan. 4 IMMUNOFLUORESCENCE LOCALIZATION OF AQUAPORIN DURING SALINITY ADAPTATION KAPPER, M.A.*; DEPAOLO, C.; Central Connecticut State Univ., New Britain kapper@ccsu.edu
During adaptation to changes in salinity osmoconforming intertidal and estuarine molluscs utilize a comprehensive suite of strategies to maintain osmotic homeostasis. Behavioral methods temporarily delay contact between cells and altered environmental salinities while physiological mechanisms modulate the concentration of intracellular free amino acids to equilibrate the concentration of intracellular and extracellular osmolytes. We are hypothesizing that during the process of physiological salinity adaptation in the ribbed mussel Geukensia demissa, the water permeability of cell membranes is modulated by trafficking aquaporin water channel proteins from the cell membrane to intracellular vesicles, effectively reducing potential water flux between the extracellular and intracellular compartments, allowing more time for adjustment of the intracellular solute concentration. We are developing immunofluorescence microscopy procedures to visualize the subcellular location of aquaporin-2 in mussel gills. Gills from mussels maintained at 15oC and 15o/ooS were excised and incubated at 15oC and 30o/ooS with aeration. Samples were fixed, embedded in paraffin and sectioned (5 microns) at 0, 30, 60, 120 and 180 minutes following salinity transfer. One set of slides was stained with hematoxylin and eosin for brightfield microscopy while a second set was exposed to an anti aquaporin 2 antibody followed by a secondary antibody conjugated to the fluorescent dye Alexa Fluor 568 and examined by fluorescence microscopy at 1000X magnification. Immunofluorescence preparations omitting the primary anti AQP antibody were used as controls, allowing tentative intracellular localization of the aquaporin protein. Supported by CSU AAUP and Student Faculty Research Grants to MAK.
