Searching of membrane target for circadian clock

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AACIMP 2011 Summer School. Neuroscience Stream. Lecture by Nikolai Kononenko.

AACIMP 2011 Summer School. Neuroscience Stream. Lecture by Nikolai Kononenko.

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  • 1. Searching of a membrane target for mammalian circadian clock responsible for circadian modulation of firing rate Nikolai I. Kononenko Department of General Physiology of Nervous System, Bogomoletz Institute of Physiology, Kiev, Ukraine
  • 2. 1. The suprachiasmatic nucleus (SCN) of the hypothalamus is the primary biological clock regulating circadian rhythms in mammals
  • 3. 2. Individual SCN neurons express self-sustained circadian oscillations. As a result of internal coupling, the SCN generates a coherent output signal
  • 4. Circadian-clock core Messenger Membrane target Hz
  • 5.
    • There are two common features for presented above hypotheses:
    • All experiments were done employing whole cell recordings
    • 2. Spike-associated currents are key targets for circadian modulation of firing rate
    • Our approach was based on on-cell (cell-attached) recordings of electrical events in SCN neurons
  • 6. Multielectrode array dish Hz 0 1 2 3 4 5 (Days) 1 min
  • 7. Spike-associated currents are key targets for circadian modulation of firing rate First of all, we asked whether spike-associated currents (i.e., currents active only during action potentials) are indeed primary membrane target(s) responsible for circadian modulation of firing rate (CMFR).
  • 8. B A Action potentials across the circadian cycle Thus, result allowed preliminarily to suggest that spike-associated currents are not a key targets for circadian modulation of firing rate.
  • 9. Nature, 2002, 16 , 286-290
  • 10. Effect of nifedipine on circadian firing rhythms in SCN neurons
  • 11. Nature Neuroscience, 2005, 8 , 650-656
  • 12. Effect of 4-aminopyridine on circadian rhythms of firing rate in SCN neurons
  • 13. A B Cd 2+ does not suppress immediately spontaneous activity and its circadian modulation (n=13 neurons)
  • 14. Spike-associated channels are not the principal determinants of circadian modulation of firing rate
  • 15. Two acutely isolated SCN neurons on the bottom of a Petri dish 6 days after isolation 100  m
  • 16. On-cell recording of single channels 2 pA 100 ms
  • 17. Spontaneous firing of isolated SCN neurons and circadian modulation of their firing rate Noon Midnight Noon of the next day
  • 18. 2 s (A) 5 s (B) 7 pA 20 mV A B 0 pA C V h = -90 mV V h = -65 mV 2 s 5 pA Fluctuation of membrane potential produces spontaneous activity of SCN neuron
  • 19. The numerous single-channel inward currents we observed during on-cell recording of spontaneous electrical firing in isolated SCN neurons led us to study the properties of the corresponding channels in their relation to spontaneous electrical firing
  • 20. 20 ms 2 pA 8 pA 20 mV A1 A2 A3 B C
  • 21. We have revealed a novel set of subthreshold, voltage-dependent cation (SVC) channels that are active at resting potential and increase their open probability with membrane depolarization
  • 22. A C B 20 pA 0.016 2 s 10 s 10 pA 0.032 -15 s 15 s 0 -100 s 0 100 s * * 2 pA 200 ms
  • 23. Correlation between Po of single-channel activity and spontaneous firing in isolated SCN neuron 1 s 10 pA 2 pA 0.06 200 ms
  • 24. Voltage dependence of persistent single channel
  • 25. We hypothesize that a sufficient number of SVC channel openings would result in threshold depolarization of the SCN neuronal membrane and spontaneous electrical firing
  • 26. A B db-cGMP application evokes spontaneous electrical activity in acutely isolated SCN neurons
  • 27.  
  • 28. g C =40 pS; E C =0 mV  o =0.2 ms;  fc =1 ms Single-channel openings (N=1)
  • 29. 36 channels ~0.017 Hz 44 channels ~3.2 Hz 55 channels ~10 Hz B A 40 mV Single-channel openings produce spontaneous firing in a model SCN neuron 1 s
  • 30. Minimal mammalian circadian clockwork model (circadian oscillator) From Scheper et al., J Neurosci 1999 Period Period Period*
  • 31. A B Circadian modulation of the number of single channels produces modulation of the action potential firing rate Circadian peaks of 3 intact SCN neurons in MED
  • 32.
    • Conclusions
    • The present findings bring together several lines of study seeking a membrane target of the circadian clock, and bridge the gap between single-channel physiology and features of circadian rhythms of firing rate.
    • The SVC single channels described here are proposed to be a key membrane target mediating the effects of circadian clock protein concentration on electrical firing rate.
    • Concomitantly, these data delineate a novel pathway that links the core circadian clock with membrane events regulating spontaneous firing in SCN neurons.
  • 33. Acknowledgments: The author thanks Dr. F.E. Dudek (University of Utah School of Medicine, Salt Lake City, USA) and Dr. S. Honma (Hokkaido University Graduate School of Medicine, Sapporo, Japan), in whose laboratories experimental data were obtained, and Dr. N.M. Berezetskaya (Institute of Physics, Kiev, Ukraine), who converted differential equations into computer programs.