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Biosensors Show the Pharmacokinetics of S-Ketamine in the Endoplasmic Reticulum

Bera, Kallol and Kamajaya, Aron and Shivange, Amol V. and Muthusamy, Anand K. and Nichols, Aaron L. and Borden, Philip M. and Grant, Stephen and Jeon, Janice and Lin, Elaine and Bishara, Ishak and Chin, Theodore M. and Cohen, Bruce N. and Kim, Charlene H. and Unger, Elizabeth K. and Tian, Lin and Marvin, Jonathan S. and Looger, Loren L. and Lester, Henry A. (2019) Biosensors Show the Pharmacokinetics of S-Ketamine in the Endoplasmic Reticulum. Frontiers in Cellular Neuroscience, 13 . Art. No. 499. ISSN 1662-5102. PMCID PMC6874132. doi:10.3389/fncel.2019.00499.

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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.

Item Type:Article
Related URLs:
URLURL TypeDescription CentralArticle
Muthusamy, Anand K.0000-0003-1041-914X
Nichols, Aaron L.0000-0001-9341-0049
Grant, Stephen0000-0003-0923-8886
Jeon, Janice0000-0002-8483-586X
Bishara, Ishak0000-0001-7563-1980
Tian, Lin0000-0001-7012-6926
Marvin, Jonathan S.0000-0003-2294-4515
Lester, Henry A.0000-0002-5470-5255
Additional Information:© 2019 Bera, Kamajaya, Shivange, Muthusamy, Nichols, Borden, Grant, Jeon, Lin, Bishara, Chin, Cohen, Kim, Unger, Tian, Marvin, Looger and Lester. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. Received: 19 August 2019; Accepted: 22 October 2019; Published: 12 November 2019. Data Availability Statement: The datasets generated for this study are available on request to the corresponding author. Author Contributions: KB, AK, AS, PB, IB, TC, AM, SG, CK and JM: performed experiments. KB, AK, AS, AM, AN, JJ, EL, BC, JM and HL: analysis. BC, JM, LL and HL: research direction. EU and LT: constructs. AN, BC, KB, EL, LL and HL: manuscript preparation and revision. LL, KB and HL: funding. This research was supported by grants from US National Institutes of Health (GM123582, MH120823, DA046122, NS090604, NS013522, MH107056), the California Institute for Regenerative Medicine (EDUC2-08398), the Brain and Behavior Research Foundation (NARSAD), the Della Martin Foundation, the Howard Hughes Medical Institute, the Caltech CI2 program, Caltech SURF donors David and Karen Rossum, and the Mistletoe Foundation. Conflict of Interest: LT is the founder of Seven Biosciences. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. Jacob P. Keller: advice on pH and tubing. Laura Luebbert: help with experiments. Luke L. Lavis: synthesis of N,N-dimethyl-S-ketamine. Michael Maher: advice on ketamine. Anindya Bhattacharya: advice on ketamine. Daniel Wagenaar: construction of LED light sources. Lauren M. Barnett: advice on photochemistry. Eric R. Schreiter: biosensors. Jonathan Wang: technical help. Margaret Jefferies and Purnima Deshpande: excellent lab management at Janelia and Caltech.
Funding AgencyGrant Number
California Institute for Regenerative Medicine (CIRM)EDUC2-08398
Brain and Behavior Research FoundationUNSPECIFIED
Della Martin FoundationUNSPECIFIED
Howard Hughes Medical Institute (HHMI)UNSPECIFIED
Caltech Innovation Initiative (CI2)UNSPECIFIED
Caltech Summer Undergraduate Research Fellowship (SURF)UNSPECIFIED
Mistletoe FoundationUNSPECIFIED
Subject Keywords:antidepressants, organelles, green fluorescent protein, protein engineering and design, periplasmic binding proteins (PBPs), inside-out pharmacology, iSketSnFR1, iSketSnFR2
PubMed Central ID:PMC6874132
Record Number:CaltechAUTHORS:20191218-090117543
Persistent URL:
Official Citation:Bera K, Kamajaya A, Shivange AV, Muthusamy AK, Nichols AL, Borden PM, Grant S, Jeon J, Lin E, Bishara I, Chin TM, Cohen BN, Kim CH, Unger EK, Tian L, Marvin JS, Looger LL and Lester HA (2019) Biosensors Show the Pharmacokinetics of S-Ketamine in the Endoplasmic Reticulum. Front. Cell. Neurosci. 13:499. doi: 10.3389/fncel.2019.00499
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:100341
Deposited By: Tony Diaz
Deposited On:18 Dec 2019 18:41
Last Modified:16 Nov 2021 17:53

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