Abstract
The target for the “rapid” (<24 h) antidepressant effects of S-ketamine is unknown, vitiating programs to rationally develop more effective rapid antidepressants. To describe a drug’s target, one must first understand the compartments entered by the drug, at all levels—the organ, the cell, and the organelle. We have, therefore, developed molecular tools to measure the subcellular, organellar pharmacokinetics of S-ketamine. The tools are genetically encoded intensity-based S-ketamine-sensing fluorescent reporters, iSKetSnFR1 and iSKetSnFR2. In solution, these biosensors respond to S-ketamine with a sensitivity, S-slope = delta(F/F0)/(delta[S-ketamine]) of 0.23 and 1.9/μM, respectively. The iSKetSnFR2 construct allows measurements at <0.3 μM S-ketamine. The iSKetSnFR1 and iSKetSnFR2 biosensors display >100-fold selectivity over other ligands tested, including R-ketamine. We targeted each of the sensors to either the plasma membrane (PM) or the endoplasmic reticulum (ER). Measurements on these biosensors expressed in Neuro2a cells and in human dopaminergic neurons differentiated from induced pluripotent stem cells (iPSCs) show that S-ketamine enters the ER within a few seconds after appearing in the external solution near the PM, then leaves as rapidly after S-ketamine is removed from the extracellular solution. In cells, S-slopes for the ER and PM-targeted sensors differ by <2-fold, indicating that the ER [S-ketamine] is less than 2-fold different from the extracellular [S-ketamine]. Organelles represent potential compartments for the engagement of S-ketamine with its antidepressant target, and potential S-ketamine targets include organellar ion channels, receptors, and transporters.
Highlights
Despite half a century of research and improvement, antidepressant drugs do not work optimally
We find that signals with the iSKetSnFR constructs have brightness similar to those of the previous iNicSnFR cpGFPbased biosensors for nicotine (Shivange et al, 2019), but the dynamic range is somewhat lower for the iKetSnFRs
In the highest-ranked results, the predicted distances between the S-ketamine N atom and the aromatic groups are too great to form a cation-π interaction of the type suggested by docking, structural, and mutational studies for the iNicSnFR series with nicotine, acetylcholine, and varenicline (Shivange et al, 2019)
Summary
Despite half a century of research and improvement, antidepressant drugs do not work optimally. Selective serotonin reuptake inhibitor antidepressants help appreciable numbers of patients, their benefits appear too slowly (2–6 weeks) after treatment has begun. Administration of a single, relatively small (subanesthetic) dose of racemic ketamine for ∼1 h partially relieves depression in
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