Intracellular pH homeostasis and serotonin-induced pH changes inCalliphorasalivary glands: the contribution of V-ATPase and carbonic anhydrase

Abstract

SUMMARY

Protein kinase A dependent and independent activation of the V-ATPase in Malpighian tubules ofAedes aegypti.

“…the larval Malpighian tubules of Drosophila hydei (Bertram and Wessing, 1994) or the larval midgut of M. sexta (Zeiske et al, 2002). Indeed, intracellular acidification has also been demonstrated in salivary glands of C. vicina upon stimulation with serotonin, cAMP, forskolin or IBMX (Schewe et al, 2008), stimuli that induce assembly and activation of the V-ATPase in this tissue. Preliminary data from intracellular pH measurements using BCECF-AM performed by us show that cAMP analogues, as well as metabolic inhibitors, acidify principal cells in Malpighian tubules of adult A. aegypti, while elevating intracellular Ca 2+ concentration does not alter the pH.…”

Intracellular pH regulation in unstimulatedCalliphorasalivary glands is Na+ dependent and requires V-ATPase activity

“…Previously, we have shown that stimulation of the gland with 5-HT induces an intracellular acidification (Schewe et al, 2008). Furthermore, salivary gland cells have been demonstrated to possess carbonic anhydrase activity, and the observed 5-HT-induced acidification is attributable to stimulated cellular respiration that masks outward-directed H + pumping by V-ATPase (Schewe et al, 2008).…”

“…Data acquisition, averaging, ratioing, display, storage and monochromator control were achieved by a program written with TestPoint programming software (Keithley). Intracellular pH was calculated from the F 490 /F 439 or F 490 /F 450 ratios by using calibration data obtained with the nigericin-K + method (Schewe et al, 2008;Thomas et al, 1979). The high-K + calibration solutions contained (in mmoll -1 ) 138 KCl, 2 CaCl 2 , 2 MgCl 2 , 3 sodium glutamate, 2.8 malic acid, 10 D-glucose, 10 TRIS and 10moll -1 nigericin.…”

“…The resting pH i in Calliphora salivary glands is 7.5±0.3 (Schewe et al, 2008). To obtain initial information about any transporters that might contribute to the maintenance of the resting pH i , we removed Na + from the bath solution, because alkalizing transporters (acid extruders) are predominantly Na + dependent.…”

“…Furthermore, salivary gland cells have been demonstrated to possess carbonic anhydrase activity, and the observed 5-HT-induced acidification is attributable to stimulated cellular respiration that masks outward-directed H + pumping by V-ATPase (Schewe et al, 2008). However, whether and to what extent V-ATPase and other intracellular pH (pH i )-regulating transporters contribute to pH i regulation in unstimulated salivary gland cells remains largely unknown.…”