Zvonko Trontelj Physics Dept., University of Ljubljana and

1. SQUID sensors a successful complementary electrophysiological instrument for imaging brain and nerve activities Zvonko Trontelj Physics Dept., University of Ljubljana and Institute for Mathematics, Physics and Mechanics, Ljubljana

2. Participating Researchers University of Ljubljana: Vojko Jazbinsek, Matjaz Slibar, Ales Stampfl, Robert Zorec PTB Berlin: Sergio Erné, Lutz Trahms, Martin Burghoff Wolfgang Mueller, Gerd Wuebbeler. FU Berlin: Gabriel Curio, Peter Aust TU Darmstadt: Gerhard Thiel, Michael Wacke Vanderbilt University: Franz Baudenbacher, Luis Fong, Jenny Holzer, John Wikswo Producers of instrumentation: Neuromag-Electa, CTF

3. Outline of talk • Introduction • On SQUID sensor • Measuring systems • Basic steps in data analysis and modelingof sources of bioelectric activity • Examples: • a)Peripheral nerve studies • b) Some examples of brain studies • Conclusions

4. Objectives: • To apply SQUID(s) in order to obtain the noninvasive information on: a) Ionic currents in electrically stimulated peripheral nerve medianus. b) Localizations of epileptical focus. c) Localization and functional information on some parts of brain cortex. • To demonstrate the pre-diagnostic capabilities of SQUID(s) • To model the intracellular curent (m. field) and its relation to AP

5. SQUID sensors • What is SQUID? • What they offer to us? • Where we can use them? • Why SQUID sensors in electrophysiology?

6. Ad 1 and Ad 2 • Superconducting QUantum Interference Device • Magnetic flux-to-voltage convertor (the most sensitive sensor for quasi dc magnet. fields m.) • Measured m. field; via Amper’s law the source • Based on 3 facts described by QM - superconductivity with Cooper pairs - C. pair tunneling - m. flux quantization

7. From Josephson jct. to closed sc. circuit

8. Dc SQUID configuration

9. Outer magnetic field is present at the SQUID

10. Ad 3 and ad Ad 4 M. flux has to be transp.to SQUID SQUID has to be in m. shielded env. High sensitivity and spatio.temp. r.

11. Multichannel SQUID system for brain studies

12. Multichannel SQUID system for brain staudies

13. We measure: Electric AP K+ anesthesia technique Magnetic field SQUID Microscope Both measurements are simultaneous Part of SQUID microscope and C.c. internodal cell holder

14. SQUID microscope prepared for the C.c. inernodal cell studies (schematically)

15. Basic steps in analysis and modelling of current sources in living matter • Distribution of ionic currents in tissue. Complicated • Direct and inverse problem • The direct problem – a unique solution • F T = ZTE FE • The inverse problem is ill-posed problem • F E = ZET FT • Simple geometry – analyt. solutions, otherweise modeling

16. Simple geometry • Single cylindrically shaped cell (1D case) • DF = 0 • Bound. cond.: Fm(z) = Fi (a,z) – Fe (a,z) • n.Ji (a,z) = n.Je(a,z)

17. Simple geometry (contin.) • From Ampere law: • Bi = Integr. [G(r,a,z – z’)Ji(a,z’)]dz • Applying the Fourier and the inverse Fourier transformations one can come from potential to mag. field and v.a.v.

18. Some methods in modeling of current sources • Current multipole expansion • Current distribution with the minimum norm estimation • Covariance method (to extract the dc component of the measured modulated magnetic field data)

19. Part of an axonor(C.Corallina intern. Cell): stimulus location and measuring points; intra-,extracell. curr.; m. field

20. The time evolution of magnetic field (vert. comp.) measured in 37 points above the C. corallina

21. Examples: • Electrically stimulated peripheral nerve medianus. Simultaneus electrical and magnetic measurements.

22. Sketch of experimental setup

23. Mag. field after stimulation at t=0, x=0: a) propagation, traces at y=30mm, x=285mm, 335mm, 385mmb) polarity reversal: x=335mm, y= -30mm and 30mm

24. Linear scan of the magnetic recordings along they-axis (a)Elec. pot. rec. simult. at y=0 and x=335mm(b)

25. Isofield pattern in the x’-y plane with 20 fT steps between two isofield lines. Crosses indicates the positins of input data points . The calculated equivalent current dipol is shown.

26. CT cross-section of the right upper arm at x=335mm as seen in the distal direction. The encircled dot at the edge of median nerve is the position of equivalent cur. dipole.

27. The model calculation of the cmpound action current

28. Results of peripheral nerve study: • The point-like current dipole is a suitable model for a simple geometry as it is in this case. • The localization of the nerve was within 2 mm (based on the CT).

29. Examples: • Determination of epileptical focus in the case of focal epilepsy

30. The flowchart of the current approach to localizing epileptic focus.  A: Time domain waveforms showing epileptic spikes. B: Spectrogram showing focal increases of spectral power.C: Magnetic source image (MSI) showing an epileptic focus.

31. Spreadnon-normal (epileptic) activity

32. The location of epileptical discharges

33. Examples: • Study of functional stimulation

34. FUNCTIONAL IMAGING: Evoked response to median nerve stimulation (not clear in the average of MEG sensors (the top overlay) .The earliest peak is from the ACG. Followed by the events in the CS. The cerebellum response is seen as well.

35. Conclusions • Magnetic measurements offer also in the world of living state valuable noninvasive information. • Both, multichannel SQUID system and SQUID microscope can be applied. The last option offers good spatial resolution. • Results from magnetic measurements can be considered as complementary to the existing electric measurements in many cases. They can be combined with different imaging modalities. • SQUID measurements provide direct information on the behavior of ionic currents. • The highest spatial resolution.

36. Thank you for your attention!

37. Examples from the world of plants: • a) Simple plant cell – Internodal cell of green algae Chara corallina

38. Our cell culture

39. Chara corallinainternodal cell

40. Multi-SQUID measuring configuration (37 channels) -schematically

41. The isofield lines representation • 3 particular time values (1.3s, 1.6s, 1.9s) after the stimulus

42. The isof. representation (at 1.4s, 2.5s, 3.6s); model. calc. of current dipol and current density along the C.c. intern. c.

43. Measured and calculated AP and B

44. Spreading of excitation along the cell: v ~ 4cm/s Conductivity: si = 1.2 W -1m -1 , sex = 0.025 W -1m -1 Length of the depolarized area: l ~ 50 mm Maximal intracellular current: Ii = 1mA Some results

45. Examples from the world of plants: b) the influence of visible light on AP and onB in Chara corallina: The chemical nature of AP obtained from the non-invasive observation (by SQUID microscope) of the intracellular current under the influence of light

46. AP as function of light exposure • Light ON 20 min • Light ON 60min Protocol of the C.c.experiment with white light illumination • Light OFF reference • Light ON 10 min

47. The influence of ilumination on the measured B and AP of electrically stimulated C.c. internodal cell

48. Model which explains the illumination experiment in the context of 2nd messenger system [Ca2+]c is altered under the influence of light/dark transitions (Miller@Sanders 1987) AP can be described by an electrically stimulated release of Ca2+ from internal store: a) the voltage depend. synthesis/breakdown of the 2nd mesenger IP3 . b) the joint action of IP3 and Ca2+ on the gating of the receptor channels, which conduct Ca2+ release from internal stores. c) modification: cells move excess Ca2+ from the cytoplasm back into internal stores by an endogeneous Ca2+ pump system (described by the Hill function. Quantitative evaluation follows the Othmer model.

49. Simulated [Ca+2]c transients in response to a single electrical stimulation

50. Some results • Assuming that the activation of the Cl- channels, that cause the depolarization, is the direct consequence of the change in [Ca2+]c , the measurementsquantitavely agree well with the model.