Advertisement

GECIquant: Semi-automated Detection and Quantification of Astrocyte Intracellular Ca2+ Signals Monitored with GCaMP6f

  • Sharmila VenugopalEmail author
  • Rahul SrinivasanEmail author
  • Baljit S. KhakhEmail author
Chapter
Part of the Springer Series in Computational Neuroscience book series (NEUROSCI)

Abstract

Astrocytes display diverse and frequent intracellular Ca2+ fluctuations that are separable by virtue of their location within the cells, their magnitude, and their duration. Recently, the study of astrocyte Ca2+ signaling has rapidly advanced by the availability of genetically encoded Ca2+ indicators (GECIs) such as GCaMP3 and GCaMP6. The systematic use of GECIs is beginning to reveal the rules for astrocyte engagement within neural circuits in brain slices and in vivo. However, the richness and high numbers of Ca2+ signals that have been observed necessitate their routine detection within the complex morphology of astrocytes. To this end, in this chapter, we describe the development and features of GECIquant software that permits the semi-automated detection and quantification of astrocyte Ca2+ signals. Biological insights afforded by the use of GECIs and GECIquant are also described.

Keywords

GECI GCaMP GECIquant Calcium Astrocytes Imaging Ca2+ microdomains Local waves Soma Territory 

Notes

Acknowledgements

The authors are supported by the NINDS (BSK, SV), NIMH (BSK), and CHDI Foundation (BSK) Awards.

References

  1. Araque A et al (2014) Gliotransmitters travel in time and space. Neuron 81:728–739CrossRefPubMedPubMedCentralGoogle Scholar
  2. Attwell D et al (2010) Glial and neuronal control of brain blood flow. Nature 468:232–243CrossRefPubMedPubMedCentralGoogle Scholar
  3. Azevedo FA et al (2009) Equal numbers of neuronal and nonneuronal cells make the human brain an isometrically scaled-up primate brain. J Comp Neurol 513:532–541CrossRefPubMedGoogle Scholar
  4. Bahney J, von Bartheld CS (2014) Validation of the isotropic fractionator: comparison with unbiased stereology and DNA extraction for quantification of glial cells. J Neurosci Methods 222:165–174CrossRefPubMedGoogle Scholar
  5. Bernardinelli Y et al (2014) Activity-dependent structural plasticity of perisynaptic astrocytic domains promotes excitatory synapse stability. Curr Biol 24:1679–1688CrossRefPubMedGoogle Scholar
  6. Berridge MJ (1998) Neuronal calcium signaling. Neuron 21:13–26CrossRefGoogle Scholar
  7. Charles AC, Merrill JE, Dirksen ER, Sanderson MJ (1991) Intercellular signaling in glial cells: calcium waves and oscillations in response to mechanical stimulation and glutamate. Neuron 6:983–992CrossRefGoogle Scholar
  8. Chen TW et al (2013) Ultrasensitive fluorescent proteins for imaging neuronal activity. Nature 499:295–300CrossRefGoogle Scholar
  9. Clapham DE (2007) Calcium signaling. Cell 131:1047–1058CrossRefPubMedPubMedCentralGoogle Scholar
  10. Cornell-Bell AH, Finkbeiner SM, Cooper MS, Smith SJ (1990) Glutamate induces calcium waves in cultured astrocytes: long-range glial signaling. Science 247:470–473CrossRefGoogle Scholar
  11. Dani JW, Chernjavsky A, Smith SJ (1992) Neuronal activity triggers calcium waves in hippocampal astrocyte networks. Neuron 8:429–440CrossRefGoogle Scholar
  12. Di Castro MA et al (2011) Local Ca2+ detection and modulation of synaptic release by astrocytes. Nat Neurosci 14:1276–1284 CrossRefGoogle Scholar
  13. Filosa JA et al (2006) Local potassium signaling couples neuronal activity to vasodilation in the brain. Nat Neurosci 9:1397–1403CrossRefPubMedGoogle Scholar
  14. Henneberger C, Papouin T, Oliet SH, Rusakov DA (2010) Long-term potentiation depends on release of D-serine from astrocytes. Nature 463:232–236 CrossRefPubMedPubMedCentralGoogle Scholar
  15. Khakh BS, McCarthy KD (2015) Astrocyte calcium signaling: from observations to functions and the challenges therein. Cold Spring Harb Perspect Biol 7:a020404CrossRefPubMedPubMedCentralGoogle Scholar
  16. Khakh BS, Sofroniew MV (2015) Diversity of astrocyte functions and phenotypes in neural circuits. Nat Neurosci 18:942–952CrossRefGoogle Scholar
  17. Li D, Agulhon C, Schmidt E, Oheim M, Ropert N (2013) New tools for investigating astrocyte-to-neuron communication. Front Cell Neurosci 7:193PubMedPubMedCentralGoogle Scholar
  18. Molotkov D, Zobova S, Arcas JM, Khiroug L (2013) Calcium-induced outgrowth of astrocytic peripheral processes requires actin binding by Profilin-1. Cell Calcium 53:338–348CrossRefPubMedGoogle Scholar
  19. Morquette P et al (2015) An astrocyte-dependent mechanism for neuronal rhythmogenesis. Nat NeurosciGoogle Scholar
  20. Nedergaard M, Verkhratsky A (2012) Artifact versus reality–how astrocytes contribute to synaptic events. Glia 60:1013–1023CrossRefGoogle Scholar
  21. Panatier A et al (2011) Astrocytes are endogenous regulators of basal transmission at central synapses. Cell 146:785–798CrossRefGoogle Scholar
  22. Perez-Alvarez A, Navarrete M, Covelo A, Martin ED, Araque A (2014) Structural and functional plasticity of astrocyte processes and dendritic spine interactions. J Neurosci 34:12738–12744CrossRefPubMedGoogle Scholar
  23. Poskanzer KE, Yuste R (2011) Astrocytic regulation of cortical UP states. Proc Natl Acad Sci U S A 108:18453–18458CrossRefPubMedPubMedCentralGoogle Scholar
  24. Ridler T, Calvard S (1978) Picture thresholding using an iterative selection method. IEEE Trans Syst Man Cybern SMC-8:630–632Google Scholar
  25. Sasaki T, Matsuki N, Ikegaya Y (2011) Action-potential modulation during axonal conduction. Science 331:599–601CrossRefPubMedGoogle Scholar
  26. Sasaki T et al (2014) Astrocyte calcium signalling orchestrates neuronal synchronization in organotypic hippocampal slices. J Physiol 592:2771–2783CrossRefPubMedPubMedCentralGoogle Scholar
  27. Schindelin J et al (2012) Fiji: an open-source platform for biological image analysis. Nat Methods 9:676–682CrossRefGoogle Scholar
  28. Schneider C, Rasband W, Eliceiri K (2012) NIH image to ImageJ: 25 years of image analysis. Nat Methods 9:671–675CrossRefGoogle Scholar
  29. Shigetomi E, Kracun S, Sofroniew MV, Khakh BS (2010) A genetically targeted optical sensor to monitor calcium signals in astrocyte processes. Nat Neurosci 13:759–766CrossRefPubMedPubMedCentralGoogle Scholar
  30. Shigetomi E, Tong X, Kwan KY, Corey DP, Khakh BS (2012) TRPA1 channels regulate astrocyte resting calcium and inhibitory synapse efficacy through GAT-3. Nat Neurosci 15:70–80CrossRefGoogle Scholar
  31. Shigetomi E, Jackson-Weaver O, Huckstepp RT, O’Dell TJ, Khakh BS (2013a) TRPA1 channels are regulators of astrocyte basal calcium levels and long-term potentiation via constitutive D-serine release. J Neurosci 33:10143–10153CrossRefPubMedPubMedCentralGoogle Scholar
  32. Shigetomi E et al (2013b) Imaging calcium microdomains within entire astrocyte territories and endfeet with GCaMPs expressed using adeno-associated viruses. J Gen Physiol 141:633–647CrossRefPubMedPubMedCentralGoogle Scholar
  33. Smith SJ (1992) Do astrocytes process neural information? Prog Brain Res 94:119–136CrossRefGoogle Scholar
  34. Smith S (1994) Neural signalling. Neuromodulatory astrocytes. Curr Biol 4:807–810CrossRefGoogle Scholar
  35. Srinivasan R et al (2015) Ca(2+) signaling in astrocytes from Ip3r2(-/-) mice in brain slices and during startle responses in vivo. Nat Neurosci 18:708–717CrossRefPubMedPubMedCentralGoogle Scholar
  36. Tong X, Shigetomi E, Looger LL, Khakh BS (2013) Genetically encoded calcium indicators and astrocyte calcium microdomains. Neuroscientist 19:274–291CrossRefPubMedGoogle Scholar
  37. Volterra A, Liaudet N, Savtchouk I (2014) Astrocyte Ca2+ signalling: an unexpected complexity. Nat Rev Neurosci 15:327–335CrossRefGoogle Scholar

Copyright information

© Springer Nature Switzerland AG 2019

Authors and Affiliations

  1. 1.Department of PhysiologyDavid Geffen School of Medicine, University of California Los AngelesLos AngelesUSA
  2. 2.Department of NeurobiologyDavid Geffen School of MedicineLos AngelesUSA
  3. 3.Department of Integrative Biology & Physiology, Division of Life SciencesUniversity of California Los AngelesLos AngelesUSA
  4. 4.Department of Neuroscience and Experimental TherapeuticsTexas A&M University College of MedicineBryanUSA

Personalised recommendations