LCIF

CIMED Live Cell Imaging Facility

New and Noteworthy:

$1000 microgrants are now available to offset the charges associated with use of the facility.  For more information see contacts above or see link to right.

1.   Overview:

The Live Cell Imaging Facility (LCIF) of the Center for Investigation of Membrane Excitability Diseases (CIMED) is focused on studying cell signaling in excitable cells with the use of optical live cell imaging methods.  We provide instruments, expertise and assistance necessary to monitor cell functions, particularly intracellular ion concentrations, with the use of fluorescent indicators.  The facility is available to researchers from inside and outside of Washington University to carry out collaborative projects related to the mission of the Center.  The facility is available to researchers interested in carrying out collaborative projects.  On January 1, 2013, we instituted a use-based repayment and grant-supported mechanisms to cover the operation costs of the facility.

2.  Equipment:

  • Two Till Photonics digital microscopes equipped with Polychrome V monochromators, high resolution (1344×1200) cooled CCD cameras, perfusion system, and controlled environment chambers.
  • A standard filter wheel based imaging rig built around a Nikon Eclipse 300 microscope with a cooled Sensicam CCD camera.
  • A dedicated IonOptix rig built around a Nikon Diaphot microscope for simultaneous analysis of sarcomere length and intracellular [Ca2+]i in isolated cardiac myocytes.
  • Olympus FV500 confocal system based on Olympus IX70 microscope with full set of Olympus lenses (4x/0.035 to 100x/1.4), which allows collecting high resolution (up to 2048×2048) images with 488 nm, 568 nm and 633 nm excitation laser lines.
  • Nikon TE2000 based TIRF system equipped with 488 nm, 531 nm and 640 nm laser lines, a dedicated Alpha TIRF (100x/1.49) lens and a fast EMCCD camera that allows single molecule FRET imaging with the use of Nikon Dual View technology.

3. Services:

  • Measurement of  intracellular free ion concentrations (Ca2+, K+, Na+, H+, etc.) and their dynamics with the use of single wavelength and ratiometric  fluorescent indicators in living cells.
  • Simultaneous measurements of cardiac myocyte contractility and [Ca2+]1 changes.
  • Analysis of cell structure, function and molecular interactions with the use of confocal, TIRF and FRET imaging.
  • Assistance in designing and performing experiments employing fluorescent indicators in living cells.

4. Staff and Contact Information:

  • Faculty Core Director:  Colin Nichols, PhD (cnichols@wustl.edu).
  • CIMED Administrator, Ms. Jayma Mikes (E-mail: jmikes@wustl.edu).
  • Core Manager:  Krzysztof (Kris) Hyrc, PhD (314-362-4876; hyrck@neuro.wustl.edu).

For all questions regarding access, training, and technical assistance please contact  Dr.  Kris Hyrc.

General inquiries should be directed to Dr. Colin Nichols or Ms. Jayma Mikes.

5. Charges:

We charge the following fees for using the facility:

Instrument

Peak Time*

Mon 9:00 AM-

Fri 5:00PM

Off-Peak Time*

Mon 5:01 PM – Fri. 9:00 PM

Nights/Weekends/Holidays*

Fri 9:01 PM –

Mon. 8:59 AM

Training fees

1-3 people

TIRF/FRET

$40/hour

$20/hour

$10/hour

$100

IonOptix

$20/hour

$10/hour

$5/hour

$50

FV500 confocal

$20/hour

$10/hour

$5/hour

$100

Till Photonics

$20/hour

$10/hour

$5/hour

$100

 *These charges apply to work done by the users themselves.  IF the work is performed by staff members, a flat fee of $50/hour applies.

PUBLICATIONS:
  • Kim E, Hyrc KL, Speck J, Salles FT, Lundberg YW, Goldberg MP, Kachar B, Warchol ME, Ornitz DM. Missense mutations in Otopetrin 1 affect subcellular localization and inhibition of purinergic signaling in vestibular supporting cells. Mol Cell Neurosci. 2011 4(6(3):655-61.
  • Crawford DC, Chang CY, Hyrc KL, Mennerick S. Calcium-independent inhibitory G-protein signaling induces persistent presynaptic muting of hippocampal synapses. J Neurosci. 2011;31(3):979-91.
  • D’Avanzo N, Hyrc K, Enkvetchakul D, Covey DF, Nichols CG. Enantioselective protein-sterol interactions mediate regulation of both prokaryotic and eukaryotic inward rectifier K+ channels by cholesterol. PLoS One. 2011 (;6(4):e19393.
  • Zeng C, Vangveravong S, Jones LA, Hyrc K, Chang KC, Xu J, Rothfuss JM, Goldberg MP, Hotchkiss RS, Mach RH Characterization and evaluation of two novel fluorescent sigma-2 receptor ligands as proliferation probes. Mol Imaging. 2011 ;10(6):420-33.
  • Toib A, Zhang HX, Broekelmann TJ, Hyrc KL, Guo Q, Chen F, Remedi MS, Nichols CG. Cardiac specific ATP-sensitive K+ channel (KATP) overexpression results in embryonic lethality. J Mol Cell Cardiol. 2012 53(3):437-45.
  • Zhang S, Hyrc K, Wang S, Wice BM. Xenin-25 increases cytosolic free calcium levels and acetylcholine release from a subset of myenteric neurons. Am J Physiol Gastrointest Liver Physiol. 2012 303(12):G1347-55.
  • Lin Z, Perez P, Sun Z, Liu JJ, Shin JH, Hyrc KL, Samways D, Egan T, Holley MC, Bao J. Reprogramming of single-cell-derived mesenchymal stem cells into hair cell-like cells. Otol Neurotol. 2012, 33(9):1648-55.
  • Wang S, Makhina EN, Masia R, Hyrc KL, Formanack ML, Nichols CG. Domain organization of the ATP-sensitive potassium channel complex examined by fluorescence resonance energy transfer. J Biol Chem. 2013 8;288(6):4378-88.
  • Zhu T, Chappel JC, Hsu FF, Turk J, Aurora R, Hyrc K, De Camilli P, Broekelmann TJ, Mecham RP, Teitelbaum SL, Zou W. Type I phosphotidylinosotol 4-phosphate 5-kinase γ regulates osteoclasts in a bifunctional manner. J Biol Chem. 2013l 288(8):5268-77.
  • Shen H, Hyrc KL, Goldberg MP. Maintaining energy homeostasis is an essential component of Wld(S)-mediated axon protection. Neurobiol Dis. 2013;59:69-79.
  • Hyrc KL, Minta A, Escamilla PR, Chan PP, Meshik XA, Goldberg MP. Synthesis and properties of Asante Calcium Red–a novel family of long excitation wavelength calcium indicators. Cell Calcium. 2013;54(4):320-33.
  • Decker CE, Yang Z, Rimer R, Park-Min KH, Macaubas C, Mellins ED, Novack DV, Faccio R. Tmem178 acts in a novel negative feedback loop targeting NFATc1 to regulate bone mass. ProcNatl Acad Sci USA 2015; 112(51):15654-9.
  • Zhang S, Wang S, Puhl MD, Jiang X, Hyrc KL, Laciny E, Wallendorf MJ, Pappan KL, Coyle JT, Wice BM. Global biochemical profiling identifies β-hydroxypyruvate as a potential mediator of type 2 diabetes in mice and humans. Diabetes. 2015; 64(4):1383-94.
  • Schindler SE, McCall JG, Yan P, Hyrc KL, Li M, Tucker CL, Lee JM, Bruchas MR, Diamond MI. Photo-activatable Cre recombinase regulates gene expression in vivo. Sci Rep. 2015; 9;5:13627.
  • Henn MC, Janjua MB, Zhang H, Kanter EM, Makepeace CM, Schuessler RB, Nichols CG, Lawton JS. Diazoxide Cardioprotection Is Independent of Adenosine Triphosphate-Sensitive Potassium Channel Kir6.1 Subunit in Response to Stress.  J Am Coll Surg. 2015; ;221(2):319-25.
  • Levin MD, Singh GK, Zhang HX, Uchida K, Kozel BA, Stein PK, Kovacs A, Westenbroek RE, Catterall WA, Grange DK, Nichols CG. K(ATP) channel gain-of-function leads to increased myocardial L-type Ca(2+) current and contractility in Cantu syndrome. Proc Natl Acad Sci USA 2016;113(24):6773-8.
  • Wakle-Prabagaran M, Lorca RA, Ma X, Stamnes SJ, Amazu C, Hsiao JJ, Karch CM, Hyrc KL, Wright ME, England SK. BKCa channel regulates calcium oscillations induced by alpha-2-macroglobulin in human myometrial smooth muscle cells.Proc Natl Acad Sci USA. 2016 ;113(16):E2335-44.
  • Mayer AL, Higgins CB, Heitmeier MR, Kraft TE, Qian X, Crowley JR, Hyrc KL, Beatty WL, Yarasheski KE, B PW, DeBosch BJ. SLC2A8 (GLUT8) is a mammalian trehalose transporter required for trehalose-induced autophagy. Sci Rep. 2016: ;(6):38586..
  • Moore BD, Jin RU, Lo H, Jung M, Wang H, Battle MA, Wollheim CB, Urano F, Mills JC. Transcriptional Regulation of X-Box-binding Protein One (XBP1) by Hepatocyte Nuclear Factor 4α (HNF4Α) Is Vital to Beta-cell Function. J Biol Chem. 2016 ;291(12):6146-57.
  • Wang S, Vafabakhsh R, Borschel WF, Ha T, Nichols CG,., Structural dynamics of potassium-channel gating revealed by single-molecule FRET. Nat Struct Mol Biol. 2016 Jan;23(1):31-36.
  • Qiao Y, Lipovsky C, Hicks S, Bhatnagar S, Li G, Khandekar A, Guzy R, Woo KV, Nichols CG, Efimov IR, Rentschler S. Transient Notch Activation Induces Long-Term Gene Expression Changes Leading to Sick Sinus Syndrome in Mice. Circ Res. 2017; 18;121(5):549-563.
  • Liu H, Javaheri A, Godar RJ, Murphy J, Ma X, Rohatgi N, Mahadevan J, Hyrc K, Saftig P, Marshall C, McDaniel ML, Remedi MS, Razani B, Urano F, Diwan A. Intermittent fasting preserves beta-cell mass in obesity-induced diabetes via the autophagy-lysosome pathway. Autophagy. 2017;13(11):1952-1968
  • Marquez-Nostra BV, Lee S, Laforest R, Vitale L, Nie X, Hyrc K, Keler T, Hawthorne T, Hoog J, Li S, Dehdashti F, Ma CX, Lapi SE. Preclinical PET imaging of glycoprotein non-metastatic melanoma B in triple negative breast cancer: feasibility of an antibody-based companion diagnostic agent. Oncotarget. 2017; 8(61):104303-104314.
  • Li G, Khandekar A, Yin T, Hicks SC, Guo Q, Takahashi K, Lipovsky CE, Brumback BD, Rao PK, Weinheimer CJ, Rentschler SL. Differential Wnt-mediated programming and arrhythmogenesis in right versus left ventricles. J Mol Cell Cardiol. 2018;123:92-107.
  • Samovski D, Dhule P, Pietka T, Jacome-Sosa M, Penrose E, Son NH, Flynn CR, Shoghi KI, Hyrc KL, Goldberg IJ, Gamazon ER, Abumrad NA. Diabetes. 2018; 67(7):1272-1284.
  • Wang S, Brettmann JB, Nichols CG. Studying Structural Dynamics of Potassium Channels by Single-Molecule FRET. Methods Mol Biol. 2018;1684:163-180.
  • Mahajan S, Decker CE, Yang Z, Veis D, Mellins ED, Faccio R. Plcγ2/Tmem178 dependent pathway in myeloid cells modulates the pathogenesis of cytokine storm syndrome. J Autoimmun. 2019. (18)30697-8.
  • Yang Z, Yan H, Dai W, Jing J, Yang Y, Mahajan S, Zhou Y, Li W, Macaubas C, Mellins ED, Shih CC, Fitzpatrick JAJ, Faccio R. Tmem178 negatively regulates store-operated calcium entry in myeloid cells via association with STIM1. J Autoimmun. 2019 1(19)30042-3.
  • Wang S, Lee SJ, Maksaev G, Fang X, Zuo C, Nichols CG. Potassium channel selectivity filter dynamics revealed by single-molecule FRET. Nat Chem Biol. 2019; (4):377-383.
  • Emfinger Cr; Lorincz R; York Natl; Singareddy S; Ikle J; Tryon R; McClenaghan C; Shyr Z, Huang Y, Reissaus C,; Meyer D, Piston D, Hyrc KL, Remedi M, Nichols CG. Adult zebrafish islets are electrically uncoupled, but exhibit KATP-dependent insulin secretion and diabetes as in mammals. Physiol Rep. 2019 Jun;7(11):e14101