Received for publication May 9, 2006.
Revised September 11, 2006.
Accepted for publication October 3, 2006.

Regulation of Human Cone CNG Channels by Endogenous Phospholipids and Exogenously Applied PIP₃

Abstract

Cyclic nucleotide gated (CNG) channels are critical components of the vertebrate visual transduction cascade involved in converting light-induced changes in intracellular cGMP concentrations into electrical signals that can be interpreted by the brain as visual information. To characterize regulatory mechanisms capable of altering the apparent ligand affinity of cone channels, we have expressed heteromeric (CNGA3 + CNGB3) human cone CNG channels in Xenopus oocytes and characterized the alterations in channel activity that occur following patch excision using patch-clamp recording in the inside-out configuration. We have found that cone channels exhibit spontaneous changes in current at sub-saturating cGMP concentrations; these changes are enhanced by application of ATP and appear to reflect alterations in channel gating. Similar to rod CNG channels, lavendustin A prevented this regulation, suggesting the involvement of a tyrosine phosphorylation event. However, the tyrosine residue in CNGB3 (Y545) that is equivalent to the critical tyrosine residues in rod and olfactory CNG channel subunits does not participate in cone channel regulation. Furthermore, the changes in ligand sensitivity of CNGA3 + CNGB3 channels were prevented by inhibition of phosphatidylinositol 3-kinase (PI3-kinase) using wortmannin or LY294002, which suggests that phospholipid metabolism can regulate the channels. Direct application of PIP₃ to the intracellular face of excised patches also resulted in down regulation of channel activity. Thus, phospholipid metabolism and exogenously applied PIP₃ can modulate heterologously expressed cone CNG channels.

Key words: Sensory (Rhodopsin, olfactory), Ion channel regulation, cAMP, cGMP