Real-Time Monitoring of Odorant Receptor Activation in Recombinant Cells

Begin with a multi-well plate containing adherent recombinant mammalian cells in a stimulation solution supplemented with luciferin, a luminescent substrate.

Incubate to allow luciferin entry to the cell.

These cells express membrane-bound odorant receptors coupled with olfactory G-proteins and receptor-transport proteins. 

Additionally, the cells express cytoplasmic Glosensor proteins, luminescent proteins sensitive to cyclic AMP. 

Add a vaporizable odorant solution in the space between the wells and place the plate within a luminometer instrument pre-equilibrated with the same odorant.

Initiate luminescence measurement.

Odorant molecules bind to the cells' odorant receptors, activating the coupled G-protein and dissociating the G-protein’s α subunit, or Gα.

The dissociated Gα subunit binds to the membrane-embedded adenylate cyclase enzyme, activating it. The activated enzyme converts the ATPs to cyclic AMPs. 

These cyclic AMPs bind with glosensor proteins, altering their conformation and enabling the luciferin binding. 

This interaction produces luminescence, confirming the activation of odorant receptors.