1. Brain energy use, control of blood flow, and the basis of BOLD signals David Attwell University College London

2. BOLD imaging Hariri et al. (2002) Science 297, 400

3. Overview Brief review of BOLD imaging Coupling of neural activity to CBF, by (i) energy use or (ii) other signalling pathways Energy budget for cerebral cortex Energy use in neuronal microcircuits: cerebellum Local regulation of CBF by glutamate Global regulation of CBF by amines Regulation of CBF by arterioles and capillaries What does BOLD measure

4. blood vessels HbO 2 Hb O2O2 FLOW basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre

5. blood vessels HbO 2 Hb O2O2 FLOW ? VOL basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre

6. Signalling from neurons to blood vessels The neuron to CBF signal is often assumed to be energy usage or energy lack (assumes CBF increases to maintain glucose/O 2 delivery to neurons) So where does the brain use energy?

8. GLUTAMATE Glial Cell 3Na + H + K + Na + Post-Synaptic Neuron Pre-Synaptic Neuron Na + Ca 2 + GLU GLN ATP 2K 3Na ATP 2K 3Na ATP 2K 3Na ATP

10. Primates vs rodents Primates: 3-10 times less cell density with same synapse density (so 3-10 times more synapses/cell) Predicts a lower overall energy usage (54% for 10-fold - experimental value is 54%) Increases fraction on glutamatergic signalling (from 34% to 74%)

12. Energy use by neuronal microcircuits: the cerebellum as an example

13. basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre

14. basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre cerebral cortexcerebellar cortex

15. basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre Predicted total ATP usage: 26.6  moles/g/min Measured: 20  moles/g/min (Sokoloff & Clarke in anaesthetized albino rats)

16. basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre Purkinje basket/stellate Golgi granule cell mossy fibre climbing fibre Bergmann ATP/sec/cell astrocyte

17. basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre Purkinje basket/stellate Golgi granule cell mossy fibre climbing fibre Bergmann ATP/sec/cell Purkinje Golgi bc/sc granule cell mossy fibre climbing fibre Bergmann astro astrocyte ATP/sec/m 2

18. ATP Usage by Subcellular Task

19. Effect of altering firing rate in a single cell type

20. Energy use by neuronal microcircuits: the cerebellum as an example (1)Most energy goes on granule cells re-mapping the sensory and motor command input arriving on the mossy fibres into a sparse coded representation used by the Purkinje cells to retrieve motor output patterns basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre

21. Energy use by neuronal microcircuits: the cerebellum as an example (1)Most energy goes on granule cells re-mapping the sensory and motor command input arriving on the mossy fibres into a sparse coded representation used by the Purkinje cells to retrieve motor output patterns (2)10 11 ATP molecules are used per second to be able to retrieve 5kB of information from each Purkinje cell (which can store 40,000 input- output associations), or 2x10 16 ATP/GB/s = (3.3x10 -8 moles/sec)x31kJ = 1mW/GB. Computer hard disks now use ~5mW/GB basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre

22. How is blood flow controlled? ML GL PC

23. Does an energy-lack signal increase blood flow? When [ATP] (or [O 2 ] or [glucose]) falls, or [CO 2 ] or [H + ] or [lactate] rises, does that make blood flow increase? In other words, do BOLD signals reflect the presence of a feedback system to conserve energy supply?

24. blood vessels HbO 2 Hb O2O2 FLOW energy lack? VOL basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre

25. What controls cerebral blood flow during brain activation? Not glucose lack (Powers et al., 1996) Not oxygen lack (Mintun et al., 2001) Not CO 2 evoked pH change (pH o goes alkaline due to CBF increase removing CO 2 : Astrup et al., 1978; Pinard et al., 1984) So CBF is not driven directly by energy lack maintaining O 2 /glucose delivery to neurons and keeping [ATP] high Powers et al., 1996

26. What controls cerebral blood flow during brain activation? CBF is not driven by energy lack Not the spike rate of principal neurons (Mathiesen et al., 1998; Lauritzen 2001) BOLD correlates (slightly!) better with synaptic field potentials than spike output (Logothetis et al., 2001) So does synaptic signalling control CBF (i.e. is it a feedforward, rather than a feedback, system)?

27. Feedforward vs feedback control of CBF Neuronal activity Energy fallsIncrease CBF Energy supplied - Negative feedback Feedforward

28. GLUTAMATE Glial Cell 3Na + H + K + Na + Post-Synaptic Neuron Pre-Synaptic Neuron + Na Ca 2 + GLU GLN ATP 2K 3Na ATP 2K 3Na ATP 2K 3Na ATP PLA 2 NOS AA,PG NO Ca 2 + PLA 2

29. Glutamate is responsible for cerebellar CBF increase Purkinje cell spikes CBF Parallel fibre stimulation Climbing fibre stimulation Matthiesen et al., 1998 CBF

30. blood vessels HbO 2 Hb O2O2 FLOW Glutamate (via neurons and glia) VOL basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre

31. Glutamate controls CBF and BOLD signals Energy calculations implicate postsynaptic currents as the main energy consumer - so if energy use drove BOLD signals, BOLD would reflect the release of glutamate In fact energy use does not drive CBF, but glutamate does - so BOLD is still likely to reflect glutamate release (via its postsynaptic actions)

32. What does BOLD measure? If BOLD signals largely reflect glutamate release: (a) BOLD does not tell us about the spike output of an area, and will only reflect principal cell activity if most glutamate is released onto principal cells (b) altered processing with no net change of glu release might produce no BOLD signal (c) altered glu release with no change of the spike output of an area could produce a BOLD signal

33. blood vessels HbO 2 Hb O2O2 FLOW Glu VOL AMINES NA, DA, 5-HT basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre

34. Control of cerebral blood flow by distributed systems using amines and ACh Dopaminergic neurons (from VTA) innervate microvessels - DA constricts (Krimer et al., 1998): D 1,2,4,5 Noradrenergic neurons (from locus coeruleus) also constrict microvessels (Raichle et al., 1975):  2 Serotoninergic neurons (from raphe) constrict cerebral arteries and microvessels (Cohen et al., 1996): 5-HT 1,2 All are wide ranging systems - control CBF globally

35. Smooth Muscle vs Pericytes blood flow capillary smooth muscle endothelial cells 10 µm SM 10 µm 5 µm s p s p pericytes

36. Smooth Muscle vs Pericytes blood flow capillary smooth muscle endothelial cells 10 µm SM 10 µm 5 µm s p s p pericytes 65% of noradrenergic innervation is of capillaries, not arterioles

37. 390s185s b 295s c d 1mM Glu 1  M NA 70s a o Peppiatt, Howarth, Auger & Attwell, unpublished Noradrenaline constricts and glutamate dilates cerebellar capillaries

38. Pericytes communicate with each other and could communicate from neurons near capillaries to precapillary arterioles

39. Implications of control of CBF by amines for neuropsychiatric imaging Clinical disorders often involve disruption of amine function (schizophrenia, Parkinson’s, ADHD) In imaging we would like a change in BOLD signals to imply an effect of the amine disorder on cortical processing If amines control CBF, altered amine function may alter the relation between neural activity and BOLD signals (extreme example: amine depletion maximally dilates vessels, so no further dilation or BOLD signal possible) Consequently altered BOLD signals may just reflect altered control of CBF, and provide no information on neural processing

40. blood vessels HbO 2 Hb O2O2 FLOW VOL AMINES NA, DA, 5-HT basket stellate granule Golgi Purkinje input mossy fibres output input climbing fibre Glu

41. BOLD imaging Hariri et al. (2002) Science 297, 400

42. Conclusions In primates, most of the brain’s energy goes on postsynaptic currents (and action potentials) CBF changes and BOLD aren’t driven by O 2 /glucose lack nor by CO 2 production, so are not driven by energy lack CBF changes and BOLD don’t correlate well with spiking Glutamate controls local CBF so BOLD signals will reflect glutamatergic signalling Amines control CBF more globally - could confound studies on amine-related diseases CONCLUSION: to interpret BOLD signals you need to consider the neural wiring of the area being studied

43. Collaborators Clare Howarth Claire Peppiatt Céline Auger Simon Laughlin