Vacuolar-type proton pump in the basolateral plasma membrane energizes ion uptake in branchial mitochondria-rich cells of killifish Fundulus heteroclitus, adapted to a low ion environment.
We examined the involvement of mitochondria-rich (MR) cells in ion uptake through gill epithelia in freshwater-adapted killifish Fundulus heteroclitus, by morphological observation of MR cells and molecular identification of the vacuolar-type proton pump (V-ATPase). MR cell morphology was compared in fish acclimated to defined freshwaters with different NaCl concentrations: low (0.1 mmol l(-1))-, mid (1 mmol l(-1))- and high (10 mmol l(-1))-NaCl environments. MR cells, mostly located on the afferent-vascular side of the gill filaments, were larger in low- and mid-NaCl environments than in the high-NaCl environment. Electron-microscopic observation revealed that the apical membrane of well-developed MR cells in low- and mid-NaCl environments was flat or slightly projecting, and equipped with microvilli to expand the surface area exposed to these environments. On the other hand, in the high-NaCl environment, the apical membrane was invaginated to form a pit, and MR cells often formed multicellular complexes with accessory cells, although the NaCl concentration was much lower than that in plasma. We cloned and sequenced a cDNA encoding the A-subunit of killifish V-ATPase. The deduced amino acid sequence showed high identity with V-ATPase A-subunits from other vertebrate species. Light-microscopic immunocytochemistry, using a homologous antibody, revealed V-ATPase-immunoreactivity in Na(+)/K(+)-ATPase-immunoreactive MR cells in low-NaCl freshwater, whereas the immunoreactivity was much weaker in higher NaCl environments. Furthermore, immuno-electron microscopy revealed V-ATPase to be located in the basolateral membrane of MR cells. These findings indicate that MR cells are the site responsible for active ion uptake in freshwater-adapted killifish, and that basolaterally located V-ATPase is involved in the Na(+) and/or Cl(-) absorbing mechanism of MR cells.
Katoh
Fumi
F
Ocean Research Institute, University of Tokyo, 1-15-1 Minamidai, Nakano, Tokyo 164-8639, Japan. fkatoh@ori.u-tokyo.ac.jp
Hyodo
Susumu
S
Kaneko
Toyoji
T
D000064:Acclimatization; D000818:Animals; D001483:Base Sequence; D001693:Biological Transport, Active; D015153:Blotting, Western; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D019085:Fluorescent Antibody Technique, Direct; D005618:Fresh Water; D023781:Fundulidae / Q000033:anatomy & histology / Q000378:metabolism*; D005880:Gills / Q000201:enzymology* / Q000648:ultrastructure; D017136:Ion Transport; D008928:Mitochondria / Q000201:enzymology*; D008969:Molecular Sequence Data; D009994:Osmolar Concentration; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism*
D017931:DNA Primers; D018076:DNA, Complementary; D025262:Vacuolar Proton-Translocating ATPases; D000254:Sodium-Potassium-Exchanging ATPase