Integrated Measurements of Electrical Activity, Oxygen Tension, Blood Flow, and Ca2+-Signaling in Rodents In Vivo

  • Claus MathiesenEmail author
  • Kirsten Thomsen
  • Martin Lauritzen
Part of the Neuromethods book series (NM, volume 90)


In order to assess perfusion and metabolic responses in relation to neural function, the cellular signaling network, including the types of neurons and astrocytes involved, and the timing of their activation need to be known/specified. Here, we present the basis for measuring brain activity and metabolism in rats and mice, which covers basic electrophysiological indicators of neuronal function, a short description of the methods commonly used for recording electrophysiological signals, examples of data analysis, and a brief look at the limitations of the methods. This chapter describes animal preparation, the origin of extracellularly recorded electrical signals, with special regard to the EEG, local field potentials, and spikes (action potentials?) in rodent preparations. We also describe methods for recording cerebral blood flow (CBF), tissue partial pressure of oxygen (tpO2), and cytosolic calcium transients. Lastly, we give examples of protocols in which electrophysiology, blood flow, cerebral rate of oxygen metabolism (CMRO2), and calcium transients have been studied together.

Key words

Synaptic activity Action potential Oxygen consumption Neurovascular coupling Cerebral blood flow 



The recent work at Lauritzen laboratory was founded by the NORDEA Foundation/Center for Healthy Aging, the Lundbeck Foundation via the Lundbeck Foundation Center for Neurovascular Signaling (LUCENS), the NOVO-Nordisk Foundation, the Danish Medical Research Council, and Foundation Leducq. We thank Micael Lønstrup for providing excellent surgical assistance; Bodil Gesslein, Barbara Lykke Lind, Krzysztof Kucharz, and Sanne Barsballe Jessen for helpful discussions.


  1. 1.
    Fish RE, Brown MJ, Danneman PJ, Karas AZ (eds) (2007) Analgesia in laboratory animals. Academic, LondonGoogle Scholar
  2. 2.
    Zhu XH, Zhang Y, Tian RX, Lei H, Zhang NY, Zhang XL et al (2002) Development of O-17 NMR approach for fast imaging of cerebral metabolic rate of oxygen in rat brain at high field. Proc Natl Acad Sci U S A 99(20):13194–13199PubMedCrossRefPubMedCentralGoogle Scholar
  3. 3.
    Zhu XH, Chen JM, Tu TW, Chen W, Song SK (2013) Simultaneous and noninvasive imaging of cerebral oxygen metabolic rate, blood flow and oxygen extraction fraction in stroke mice. Neuroimage 64:437–447PubMedCrossRefPubMedCentralGoogle Scholar
  4. 4.
    Shih AY, Driscoll JD, Drew PJ, Nishimura N, Schaffer CB, Kleinfeld D (2012) Two-photon microscopy as a tool to study blood flow and neurovascular coupling in the rodent brain. J Cereb Blood Flow Metab 32(7):1277–1309PubMedCrossRefPubMedCentralGoogle Scholar
  5. 5.
    Shih AY, Mateo C, Drew PJ, Tsai PS, Kleinfeld D (2012) A polished and reinforced thinned-skull window for long-term imaging of the mouse brain. J Vis Exp 61:e3742Google Scholar
  6. 6.
    Marker DF, Tremblay ME, Lu SM, Majewska AK, Gelbard HA (2010) A thin-skull window technique for chronic two-photon in vivo imaging of murine microglia in models of neuroinflammation. J Vis Exp 43:4059Google Scholar
  7. 7.
    Drew PJ, Shih AY, Driscoll JD, Knutsen PM, Blinder P, Davalos D et al (2010) Chronic optical access through a polished and reinforced thinned skull. Nat Methods 7(12):981–984PubMedCrossRefPubMedCentralGoogle Scholar
  8. 8.
    Kazama K, Anrather J, Zhou P, Girouard H, Frys K, Milner TA et al (2004) Angiotensin II impairs neurovascular coupling in neocortex through NADPH oxidase-derived radicals. Circ Res 95(10):1019–1026PubMedCrossRefGoogle Scholar
  9. 9.
    Nedergaard M, Ransom B, Goldman SA (2003) New roles for astrocytes: redefining the functional architecture of the brain. Trends Neurosci 26(10):523–530PubMedCrossRefGoogle Scholar
  10. 10.
    Ransom B, Behar T, Nedergaard M (2003) New roles for astrocytes (stars at last). Trends Neurosci 26(10):520–522PubMedCrossRefGoogle Scholar
  11. 11.
    Simard M, Arcuino G, Takano T, Liu QS, Nedergaard M (2003) Signaling at the gliovascular interface. J Neurosci 23(27):9254–9262PubMedGoogle Scholar
  12. 12.
    Ludwin SK (1981) Pathology of demyelination and remyelination. Adv Neurol 31:123–168, Epub 1981/01/01PubMedGoogle Scholar
  13. 13.
    Jones LL, Banati RB, Graeber MB, Bonfanti L, Raivich G, Kreutzberg GW (1997) Population control of microglia: does apoptosis play a role? J Neurocytol 26(11):755–770, Epub 1998/01/13PubMedCrossRefGoogle Scholar
  14. 14.
    Seigneur J, Kroeger D, Nita DA, Amzica F (2006) Cholinergic action on cortical glial cells in vivo. Cereb Cortex 16(5):655–668, Epub 2005/08/12PubMedCrossRefGoogle Scholar
  15. 15.
    Hess G, Gustafsson B (1990) Changes in field excitatory postsynaptic potential shape induced by tetanization in the CA1 region of the guinea-pig hippocampal slice. Neuroscience 37(1):61–69, Epub 1990/01/01PubMedCrossRefGoogle Scholar
  16. 16.
    Andersen P, Bliss TV, Skrede KK (1971) Unit analysis of hippocampal polulation spikes. Exp Brain Res 13(2):208–221, Epub 1971/01/01PubMedGoogle Scholar
  17. 17.
    Nicholson C, Llinas R (1975) Real time current source analysis using multielectrode array in cat cerebellum. Brain Res 100:418–424PubMedCrossRefGoogle Scholar
  18. 18.
    Nicholson C, Freeman JA (1975) Theory of current source-density analysis and determination of conductivity tensor for anuran cerebellum. J Neurophysiol 38(2):356–368PubMedGoogle Scholar
  19. 19.
    Nakagawa H, Matsumoto N (1998) ON and OFF channels of the frog optic tectum revealed by current source density analysis. J Neurophysiol 80(4):1886–1899PubMedGoogle Scholar
  20. 20.
    Mathiesen C, Caesar K, Thomsen K, Hoogland TM, Witgen BM, Brazhe A et al (2011) Activity-dependent increases in local oxygen consumption correlate with postsynaptic currents in the mouse cerebellum in vivo. J Neurosci 31(50):18327–18337PubMedCrossRefGoogle Scholar
  21. 21.
    Grace AA, Bunney BS (1983) Intracellular and extracellular electrophysiology of nigral dopaminergic neurons – 1. Identification and characterization. Neuroscience 10(2):301–315, Epub 1983/10/01PubMedCrossRefGoogle Scholar
  22. 22.
    Herrik KF, Christophersen P, Shepard PD (2010) Pharmacological modulation of the gating properties of small conductance Ca2+-activated K+ channels alters the firing pattern of dopamine neurons in vivo. J Neurophysiol 104(3):1726–1735, Epub 2010/07/28PubMedCrossRefGoogle Scholar
  23. 23.
    Mathiesen C, Caesar K, Akgoren N, Lauritzen M (1998) Modification of activity-dependent increases of cerebral blood flow by excitatory synaptic activity and spikes in rat cerebellar cortex. J Physiol 512(Pt 2):555–566PubMedCrossRefPubMedCentralGoogle Scholar
  24. 24.
    Harris KD, Henze DA, Csicsvari J, Hirase H, Buzsaki G (2000) Accuracy of tetrode spike separation as determined by simultaneous intracellular and extracellular measurements. J Neurophysiol 84(1):401–414, Epub 2000/07/19PubMedGoogle Scholar
  25. 25.
    Emondi AA, Rebrik SP, Kurgansky AV, Miller KD (2004) Tracking neurons recorded from tetrodes across time. J Neurosci Methods 135(1–2):95–105, Epub 2004/03/17PubMedCrossRefGoogle Scholar
  26. 26.
    Chakrabarti S, Hebert P, Wolf MT, Campos M, Burdick JW, Gail A (2012) Expert-like performance of an autonomous spike tracking algorithm in isolating and maintaining single units in the macaque cortex. J Neurosci Methods 205(1):72–85, Epub 2012/01/10PubMedCrossRefGoogle Scholar
  27. 27.
    Revsbech NP (1989) An oxygen microsensor with a guard cathode. Limnol Oceanogr 34:474–478CrossRefGoogle Scholar
  28. 28.
    Skarphedinsson JO, Harding H, Thoren P (1988) Repeated measurements of cerebral blood flow in rats. Comparisons between the hydrogen clearance method and laser Doppler flowmetry. Acta Physiol Scand 134:133–142PubMedCrossRefGoogle Scholar
  29. 29.
    Dirnagl U, Kaplan B, Jacewicz M, Pulsinelli W (1989) Continuous measurement of cerebral cortical blood flow by laser-Doppler flowmetry in a rat stroke model. J Cereb Blood Flow Metab 9(5):589–596PubMedCrossRefGoogle Scholar
  30. 30.
    Fabricius M, Lauritzen M (1996) Laser-Doppler evaluation of rat brain microcirculation: comparison with the [14C]-iodoantipyrine method suggests discordance during cerebral blood flow increases. J Cereb Blood Flow Metab 16(1):156–161PubMedCrossRefGoogle Scholar
  31. 31.
    Lauritzen M, Fabricius M (1995) Real time laser-Doppler perfusion imaging of cortical spreading depression in rat neocortex. Neuroreport 6(9):1271–1273PubMedCrossRefGoogle Scholar
  32. 32.
    Gjedde A (2005) Blood–brain transfer and metabolism of oxygen. In: Dermietzel RS, Nedergaard M (eds) Blood–brain barriers: from ontogeny to artificial interfaces. Wiley, Hoboken, NJGoogle Scholar
  33. 33.
    Caesar K, Hashemi P, Douhou A, Bonvento G, Boutelle MG, Walls AB et al (2008) Glutamate receptor-dependent increments in lactate, glucose and oxygen metabolism evoked in rat cerebellum in vivo. J Physiol 586(5):1337–1349PubMedCrossRefPubMedCentralGoogle Scholar
  34. 34.
    Caesar K, Offenhauser N, Lauritzen M (2008) Gamma-aminobutyric acid modulates local brain oxygen consumption and blood flow in rat cerebellar cortex. J Cereb Blood Flow Metab 28(5):906–915PubMedCrossRefGoogle Scholar
  35. 35.
    Piilgaard H, Lauritzen M (2009) Persistent increase in oxygen consumption and impaired neurovascular coupling after spreading depression in rat neocortex. J Cereb Blood Flow Metab 29(9):1517–1527PubMedCrossRefGoogle Scholar
  36. 36.
    Piilgaard H, Witgen BM, Rasmussen P, Lauritzen M (2011) Cyclosporine A, FK506, and NIM811 ameliorate prolonged CBF reduction and impaired neurovascular coupling after cortical spreading depression. J Cereb Blood Flow Metab 31:1588–1598PubMedCrossRefPubMedCentralGoogle Scholar
  37. 37.
    Thomsen K, Piilgaard H, Gjedde A, Bonvento G, Lauritzen M (2009) Principal cell spiking, postsynaptic excitation, and oxygen consumption in the rat cerebellar cortex. J Neurophysiol 102(3):1503–1512PubMedCrossRefGoogle Scholar
  38. 38.
    Lauritzen M (2005) Opinion: reading vascular changes in brain imaging: is dendritic calcium the key? Nat Rev Neurosci 6(1):77–85PubMedCrossRefGoogle Scholar
  39. 39.
    Jespersen SN, Ostergaard L (2012) The roles of cerebral blood flow, capillary transit time heterogeneity, and oxygen tension in brain oxygenation and metabolism. J Cereb Blood Flow Metab 32(2):264–277PubMedCrossRefPubMedCentralGoogle Scholar
  40. 40.
    Fabricius M, Akgoren N, Dirnagl U, Lauritzen M (1997) Laminar analysis of cerebral blood flow in cortex of rats by laser-Doppler flowmetry: a pilot study. J Cereb Blood Flow Metab 17(12):1326–1336PubMedCrossRefGoogle Scholar
  41. 41.
    Akgoren N, Mathiesen C, Rubin I, Lauritzen M (1997) Laminar analysis of activity-dependent increases of CBF in rat cerebellar cortex: dependence on synaptic strength. Am J Physiol 273(3 Pt 2):H1166–H1176PubMedGoogle Scholar
  42. 42.
    Caesar K, Gold L, Lauritzen M (2003) Context sensitivity of activity-dependent increases in cerebral blood flow. Proc Natl Acad Sci 100(7):4239PubMedCrossRefPubMedCentralGoogle Scholar
  43. 43.
    Nimmerjahn A, Kirchhoff F, Kerr JND, Helmchen F (2004) Sulforhodamine 101 as a specific marker of astroglia in the neocortex in vivo. Nat Methods 1(1):31–37PubMedCrossRefGoogle Scholar
  44. 44.
    Wang Q, Shui B, Kotlikoff MI, Sondermann H (2008) Structural basis for calcium sensing by GCaMP2. Structure 16(12):1817–1827, Epub 2008/12/17PubMedCrossRefPubMedCentralGoogle Scholar
  45. 45.
    Logothetis NK, Pauls J, Augath M, Trinath T, Oeltermann A (2001) Neurophysiological investigation of the basis of the fMRI signal. Nature 412(6843):150–157PubMedCrossRefGoogle Scholar
  46. 46.
    Kayser C, Kim M, Ugurbil K, Kim DS, Konig P (2004) A comparison of hemodynamic and neural responses in cat visual cortex using complex stimuli. Cereb Cortex 14(8):881–891PubMedCrossRefGoogle Scholar
  47. 47.
    Thomsen K, Offenhauser N, Lauritzen M (2004) Principal neuron spiking: neither necessary nor sufficient for cerebral blood flow in rat cerebellum. J Physiol 560(Pt 1):181–189PubMedCrossRefPubMedCentralGoogle Scholar
  48. 48.
    Gobel W, Helmchen F (2007) New angles on neuronal dendrites in vivo. J Neurophysiol 98(6):3770–3779, Epub 2007/09/28PubMedCrossRefGoogle Scholar
  49. 49.
    Yang G, Huard JM, Beitz AJ, Ross ME, Iadecola C (2000) Stellate neurons mediate functional hyperemia in the cerebellar molecular layer. J Neurosci 20(18):6968–6973PubMedGoogle Scholar
  50. 50.
    Gold L, Lauritzen M (2002) Neuronal deactivation explains decreased cerebellar blood flow in response to focal cerebral ischemia or suppressed neocortical function. Proc Natl Acad Sci U S A 99(11):7699–7704PubMedCrossRefPubMedCentralGoogle Scholar
  51. 51.
    Caesar K, Thomsen K, Lauritzen M (2003) Dissociation of spikes, synaptic activity, and activity-dependent increments in rat cerebellar blood flow by tonic synaptic inhibition. Proc Natl Acad Sci U S A 100(26):16000–16005PubMedCrossRefPubMedCentralGoogle Scholar
  52. 52.
    Attwell D, Laughlin SB (2001) An energy budget for signaling in the grey matter of the brain. J Cereb Blood Flow Metab 21(10):1133–1145PubMedCrossRefGoogle Scholar
  53. 53.
    Lauritzen M (2001) Relationship of spikes, synaptic activity, and local changes of cerebral blood flow. J Cereb Blood Flow Metab 21(12):1367–1383PubMedCrossRefGoogle Scholar
  54. 54.
    Southam E, Morris R, Garthwaite J (1992) Sources and targets of nitric oxide in rat cerebellum. Neurosci Lett 137:241–244PubMedCrossRefGoogle Scholar
  55. 55.
    Offenhauser N, Thomsen K, Caesar K, Lauritzen M (2005) Activity-induced tissue oxygenation changes in rat cerebellar cortex: interplay of postsynaptic activation and blood flow. J Physiol (Lond) 565(1):279–294CrossRefGoogle Scholar
  56. 56.
    Leithner C, Royl G, Offenhauser N, Fuchtemeier M, Kohl-Bareis M, Villringer A et al (2010) Pharmacological uncoupling of activation induced increases in CBF and CMRO2. J Cereb Blood Flow Metab 30(2):311–322, Epub 2009/10/02PubMedCrossRefPubMedCentralGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Claus Mathiesen
    • 1
    Email author
  • Kirsten Thomsen
    • 1
    • 2
  • Martin Lauritzen
    • 1
    • 2
  1. 1.Department of Neuroscience and Pharmacology, Faculty of Health and Medical Sciences, Center for Healthy AgingUniversity of CopenhagenCopenhagenDenmark
  2. 2.Department of Clinical NeurophysiologyGlostrup HospitalGlostrupDenmark

Personalised recommendations