P-gp-induced modulation of regulatory volume increase occurs via PKC in mouse proximal tubule

Area of Science:

• Renal Physiology
• Cell Biology
• Molecular Pharmacology

Background:

• P-glycoprotein (P-gp) is an efflux transporter involved in various physiological processes.
• Regulatory Volume Increase (RVI) is a critical cellular response to hyperosmotic stress.
• The role of Protein Kinase C (PKC) in P-gp-mediated RVI remains incompletely understood.

Purpose of the Study:

• To investigate the involvement of PKC in P-gp-modulated RVI in mouse proximal tubule S2 segments.
• To elucidate the signaling pathways downstream of PKC in this process.

Main Methods:

• Utilized isolated proximal tubule S2 segments from wild-type (WT) and mdr1a/mdr1b gene knockout (KO) mice.
• Applied hyperosmotic solutions with or without P-gp activators/inhibitors (mannitol, phorbol 12-myristate 13-acetate [PMA], verapamil, cyclosporin A).
• Investigated the role of PKC inhibitors (staurosporine, calphostin C), cytoskeletal disruptors (colchicine, cytochalasin B), and PI 3-kinase blockers (wortmannin, LY-294002).

Main Results:

• Hyperosmotic conditions activated PKC and elicited RVI in KO tubules, but not WT tubules.
• PMA activated PKC and elicited RVI in WT tubules, an effect abolished by PKC inhibitors.
• P-gp inhibitors induced RVI in WT tubules, an effect dependent on PKC.
• PKC inhibitors abolished RVI in KO tubules, while PMA did not restore it.
• PMA-induced RVI in WT tubules and RVI in KO tubules were sensitive to microtubule and microfilament disruption, and wortmannin, but not LY-294002.

Conclusions:

• P-gp modulates RVI in mouse proximal tubules through PKC activation.
• Microtubules, microfilaments, and a wortmannin-sensitive, LY-294002-insensitive PI 3-kinase pathway contribute to PKC-induced RVI.