sorting the connections with multi-electrode neuronal ensemble recording techniques:
DESCRIPTION
Neuronal Networks Laboratory. Sorting the connections with multi-electrode neuronal ensemble recording techniques: Single-unit and local field potential activity from rat to man Dr Rob Mason Institute of Neuroscience School of Biomedical Sciences - PowerPoint PPT PresentationTRANSCRIPT
Sorting the connections with multi-electrode neuronal ensemble recording
techniques:
Single-unit and local field potential activity from rat to man
Dr Rob Mason
Institute of Neuroscience
School of Biomedical Sciences
University of Nottingham Medical School
and the LAB TEAM
Neuronal Networks Laboratory
LAB TEAM
epilepsy Ben Coomber [Dr Mike O’Donoghue ~ Dept
Neurology, QMC]
Clare Roe schizophrenia Dr Jill Suckling [Prof CA Marsden]
Dr Dissanayake anxiety Dr Carl Stevenson [Prof CA Marsden]
pain Dr Steve Elmes [Dr V Chapman]
rodent USVs Beth Tunstall [Dr S Beckett]
data analysis Margarita Zachariou [Prof S Coombes / Dr M Owen]
[Mathematics Dept]
Dr David Halliday (University of York)Prof D Auer (fMRI / phMRI ~ QMC)
Neuronal Networks Laboratory
Seminar overview
• Multichannel Electrophysiological Recording Technologies
Illustrate with reference to 2 experimental rodent model projects:
• Hippocampus-mPFC ~ Epilepsy model – role of endocannabinoid system
• Sensory gating in hippocampus & mPFC ~ Schizophrenia model – PCP effects
• Amygdala-mPFC ~ Stress models – maternal separation & “drug stressors”• Left-Right mPFC ~
• Rat language ~ ultrasound vocalisations (USVs) & affective state
Neuronal Networks Laboratory
Sorting the connections with multi-electrode neuronal ensemble recording techniques
To & From ~ rat prefrontal cortex
Sorting signal from noise
Ensemble Neuronal Unit activity
LFP activity
DATA Analysis & Interpretation
Distribution of neurones contributing to signals recorded by tetrode array
after Busaki 2004
Bionics 100-channel (“hedgehog”) array
Michigan MEAprobe 16-channels
NBLabs 8-channel array
4-channel - independent manipulation
Examples of Electrode Arrays
MULTIPLE ELECTRODE ARRAY RECORDING in vivo
• Single-site recording
e.g. hippocampal sub regions (CA1 & CA3)
• Multiple-site recording
e.g. prefrontal cortex & hippocampus
NBLabs 16-channel micro-wire array
• CA1 & CA3
Multiple Electrode Recordings in vivosimultaneous multichannel spike & field potential
recording
• 64-channel MAP system• 32-channel MAP system – with CinePlex for behavioural studies • 64-chanel Recorder system• 16-chanel Recorder system used MAP & Recorder for in vitro MEA studies with brain slices & neuronal cultures
Extracellular Recording - signal filtering & discrimination
AP Spike discrimination: separate action potential (AP) signal from noise
• AP amplitude detection / AP waveform shape recognition
Signal filtering: separate unit activity and Local Field Potentials (LFPs)
0 0.10 0.20 TIME (s)
Raw
Sig
nal
(V
) 0
100
100
0Filt
ered
Sig
nal
(V
)
Impulse Events
Units + EEG / LFP
Units
Multiple Neuronal (spike) Recording - two electrodes
micro-electrodes
Neurones
Amplifiernerve impulses
(action potential “spikes”)
Electrode 1
Electrode 2
Off Line Sorting of spike waveforms - single unit isolation
Movie: Sorting of unit spike data using Principal Component Analysis
• distinct AP spike waveforms represented as clusters in 3D space• 7 units isolated – each unit colour-coded
Multiple (ensemble) neurone recording
Advantages
• Investigating neuronal ensemble/network function - closer to working “brain”
• Good experimental design - fewer animals required (“3Rs” ~ Home Office)
• Masses of data
Disadvantages
• Masses of data? Data processing? Data interpretation
Data Visualisation Emergent Properties
Population Dynamics
Unit activity
Single units – spike rasters /FRH / ISIH / PSTHs burst analysis /
Unit pairs – cross-correlation / coherence /Unit ensembles – PCA / ICA / synchrony index / PDC
Local Field Potentials
FFT / spectrogramsLFP/EEG signal bandsLFP-unit coherenceLFP PDC
ENSEMBLE DATA - DISPLAY & ANALYSIS
MASSIVE data sets
• Dual/Triple site recordings ~ 64 channels simultaneous units & LFPs
VTA - mPFC hippocampus - mPFCamygdala - mPFCmPFC – mPFCspinal cord – thalamus – cerebral cortex
• Systemic pharmacological manipulation
• Local pharmacological manipulation
“injectrode” integrated microiontophoresis with recording array
• Electrical stimulation – periphery / CNS structures
• Independent electrode array microdrive – 8 channel drive
• Anaesthetised & awake-behaving preparations
Neuronal Networks Laboratory
LAB EXPERIMENTAL DIRECTIONS
mPFCx interconnectivity & functional context – in vivo studies
maternal separation - Depression- Stress
circadian - affective states
- Sensory Gating- Schizophrenia- Epilepsy
USVs- affective states
medial Pre-Frontal Cortex
Amygdalanucleus AccumbensHippocampusSCN
Contralateral
mPFC
VTA
schizophreniadrugs of abuse
PVt
bladder
University of Nottingham Medical SchoolNeuronal Networks Laboratory
Project #1: Epilepsy
• kainate-induced epileptiform activity ~ TLE
• functional network interactions ~ hippocampus mPFC
• role of endocannabinoid system ~ CB1R pharmacology
• perforant path stimulation-evoked seizure activity
Neuronal Networks Laboratory
FAAHMGL
Reuptake
Endo-CANNABINOID System
• Endogenous cannabinoids (eCBs) identified
e.g. anandamide (AEA) and 2-arachidonylglycerol (2-AG)
• Act at cannabinoid G protein coupled-Receptors: CB1 & CB2 - ? CB3
• eCBs are synthesised post-synaptically on-demand.
• eCBs act at pre-synaptic CB1 receptors.
• eCB reuptake occurs via a transporter.
• Metabolism:– AEA by fatty acid amide
hydrolase (FAAH)– 2-AG by monoacylglycerol
lipase (MGL).Figure taken from Wilson & Nicoll (2002) Science.
• Kainic acid (KA): established convulsive agent producing seizures in awake rats ~ targeting temporal lobe
Model uses KA, administered systemically (10mg/kg, i.p.) anaesthetized rats ~ ensemble neuronal unit and LFP activity
• Study aims to establish whether URB597, selective inhibitor of FAAH enzyme (eCB levels), attenuates KA-evoked neuronal activity
• Role of CB1 cannabinoid receptors assessed using selective CB1 antagonist AM251
Epilepsy Study: AIMS
Anandamide Metabolism by FAAH
URB597
Inhibits anandamide metabolism
KA + Vehicle: Rat #1
Basal: Rat #1 Basal: Rat #2
KA + URB597: Rat #2
Unit and LFP activity in mPFC and hippocampus
0 20 40 60 80 100 1200
200
400
600
800
1000KA + VehicleKA + URB587 (0.3)KA + URB597 (1.0)
Time (min)
% o
f b
asal
fir
ing
rat
e
• Effects on hippocampal neural firing rate (~40 cells; n=5 rats)
-100 0 100 200 300 400 500
Time (ms)
-0.3
-0.2
-0.1
-0
0.1
Basalepoch_30minepoch_40minepoch_50minepoch_60minepoch_90minepoch_120min
Spike-triggered averaging of LFP using 1a as ref cell
LF
P A
mp
litu
de
(m
V)
• Effects on spike-triggered averaging of mPFC LFPs
Post-KA administration
-1 -0.5 0 0.5 10
2
4
6
8
10
12
14
sig001a
-1 -0.5 0 0.5 10
20
40
60
sig006a
-1 -0.5 0 0.5 1Time (sec)
0
2
4
6
8
sig008a
-1 -0.5 0 0.5 10
2
4
6
8
sig022e
-1 -0.5 0 0.5 10
2
4
6
8
sig023a
-1 -0.5 0 0.5 1Time (sec)
5
10
15
sig024f
Crosscorrelograms, reference = sig006a, bin = 1 ms
Co
un
ts/b
in
Ref unit
n=2
n=12
n=7
Cross-correlation analysis ~ PFC – PFC / PFC- Hippocampal neuronal pairs
# units
Cross-correlogram TF #1
+1s 0 -1s
Time (s)
Time Frame #1-13Time (s)
Time-Series cross-correlograms – TF #1-13
corr
elat
ion
str
eng
th
Cross-correlation analysis of PFC neuronal pairs
• Effect of kainate (10mg/kg; i.p.) administration at epoch 4
Time Frames #1-13
corr
elat
ion
str
eng
th
PDC Analysis – unit ensemble data
• PDC was applied to identify the direction of activity between hippocampusand mPFC - technique that has the potential to reveal the neuronal ensemble drives.
Note: the magnitude of the classical coherence gives no information about directional connectivity - but its phase may do so.
University of Nottingham Medical SchoolNeuronal Networks Laboratory
Project #2: Schizophrenia
• auditory-evoked sensory gating ~ hippocampus & mPFC
• effects of PCP / ketamine ~ model
• effects of social isolation ~ model
Neuronal Networks Laboratory
Sensory gating in hippocampus:
A model for schizophrenia ?
• Sensory gating: mechanism(s) by which irrelevant sensory information is filtered ~ enables efficient information processing
• Auditory Conditioning-Test paradigm: measures reduction in auditory-evoked response produced by Test stimulus following a Conditioning stimulus
• Stimuli: 3kHz sine-wave / 10ms duration / presented 500ms apart / 80-90dB
• human P50 wave = rat N40 component
• Gating absent in - schizophrenic patients (& family)- normal volunteers given PCP /
amphetamine
- rats given PCP / amphetamine
Neuronal Networks Laboratory
1sLFP2 averaged 128 trials
Units
LFPs
Hippocampal CA3 - auditory-evoked unit & LFP activity
- 128
- 1
Trial #
sCs Ts
mV
N40N40
Averaged LFP
T/C ratio = 55%
eventtone
dentate gyrus CA3 region CA3 region
Single-unit PSTHs ‘ rasters histograms – “gating rats”
Unit 1
Unit 2
Unit 3
Unit 4
Spike raster
PSTH
LFP
LFP trial raster
Hippocampal CA3 auditory-evoked unit & LFP activity
Effects of PCP (1mg/kg i.p.) attenuates / abolishes sensory gating
Basal [128 trials]
T/C ratio = 32%i.e. exhibits gating
45mins after PCP
T/C ratio = 66%gating attenuated
1
Trail #
128
SUMMARY III
Sensory Gating Studies
LFP studies:
• Demonstrate sensory gating in isoflurane-anaesthetised rat : T/C ratio = 35 ± 15%
• SG is abolished / attenuated following PCP : T/C ratio = 65 ± 5%
• control rats SG is unaffected by clozapine : T/C ratio 40%
• Clozapine (5mg.kg-1) blocks action of PCP on SG : T/C ratio 35%
Unit studies:
• Similar observations
Project #3:
Rat ultrasound vocalisations (USVs) & affective state
• Sorting the pips from the squeaks
•Role of nuc Accumbens in USV-mediated Behaviours ~50 kHz (Reward) call(Brudzynski, 2001)
• nucleus Accumbens - mPFCx functional connectivity
Neuronal Networks Laboratory
-68
-63
-59
-54
-49
-45
-40
-34
dB20 ms
25
50
75
100
125
kHz
1V
CURRENT APPROACHES & FUTURE DIRECTIONS
Human Studies
Neuro-Robotics & NeuroProsthetics
Hybrid Brain-Machine interfaces (HBMIs)
• Cochlear implants
• Monitoring & control of epileptic seizures
• Robotic limbs
Neuronal Networks Laboratory
(A) Seizure control
(B) Robotic arm control
Chips in the Brain ~ Brain-Machine Interfaces (BMIs) ~ Brain-Computer Interfaces
Current issue (May 2007) Scientific American MIND 18 (2): 65-69
Unit activity
vs
LFPs
Signal choice &
Algorithm
Brain-Computer Interfaces
Brain-Computer Interfaces
Recording WorkStation – Plexon IncElectrode array Implant
(1) ENSEMBLE RECORDINGS OF HUMAN SUBCORTICAL NEURONS AS A SOURCE OF MOTOR CONTROL SIGNALS FOR A BRAIN-MACHINE INTERFACE
Parag G. Patil et al - Neurosurgery (2004) – Duke University
~ 32-channel PtIr 40m wire array
~ 4 Deep Brain Stimulation electrodes
[Medtronics DBS]
Unit recordings – 4 microwires
Thalamic VOP/VIN STN
[Plexon Inc MAP recording system]
(2) ENSEMBLE RECORDINGS OF HUMAN SUBCORTICAL NEURONS AS A SOURCE OF MOTOR CONTROL SIGNALS FOR A BRAIN-MACHINE INTERFACE
Parag G. Patil et al - Neurosurgery (2004) – Duke University
STN recording ~ 24 units
Patient performance motor task
REFERENCES Human / Primate Recording Reviews
• LEARNING TO CONTROL A BRAIN–MACHINE INTERFACE FOR REACHING AND GRASPING BY PRIMATES
JM Carmena, et alPLoS Biology ~ http://biology.plosjournals.org 1(2): 193-208 (2003)
• ENSEMBLE RECORDINGS OF HUMAN SUBCORTICAL NEURONS AS A SOURCE OF MOTOR CONTROL SIGNALS FOR A BRAIN-MACHINE INTERFACE
PG Patil, JM. Carmena, Miguel AL Nicolelis & DA TurnerNeurosurgery 55(1): 27-38 (2004)
• ASSISTIVE TECHNOLOGY & ROBITC CONTROL USING MOTOR CORTEX ENSEMBLE-BASED NEURAL INTERFACE SYSTEMS IN HUMANS WITH TETRAPLEGIA
JP Donoghue et alJ . Physiol 579(3) 603-611 (2007)
University of Nottingham Medical SchoolNeuronal Networks Laboratory
Lab web sitewww.nottingham.ac.uk/neuronal-networks
Neuronal Networks Laboratory
That’s all folks
in vivo Basal
Bicuculline (7.5 mg.kg-1 i.v.)
in vivo Basal
Kainate (10 mg.kg- I i.v.)
University of Nottingham Medical SchoolNeuronal Networks Laboratory
Project #4: Affective state
Cortico-limibic network interactions
• anxiety effects of maternal separationpharmacologically-induced (e.g. FG-7142)
anxiety
• behavioural sequalaerodent ultrasound vocalisations
Neuronal Networks Laboratory
University of Nottingham Medical SchoolNeuronal Networks Laboratory
Project #5:
Nociception & pain management
• dual spinal cord / supraspinal recording
Role of endocannabinoid system in normal physiology and pain (e.g. neuropathic) states
Neuronal Networks Laboratory
-2 0 2 4 6Time (sec)
10
20
30
40
sig004a
Perievent Rasters, reference = Event002, bin = 100 ms
Fre
quen
cy (
imp/
sec)
innocuous (7g) stimulation
-2 0 2 4 6Time (sec)
20
40
60
sig004a
Perievent Rasters, reference = Event002, bin = 100 ms
Fre
quen
cy (
imp/
sec)
noxious (65g) stimulation
Mechanically-evoked response in somatosensory thalamus (VPM)
• Cannabinoid receptor agonists are antinociceptive.
• CB1 predominantly expressed in the CNS but also present in the periphery.
• CB2 agonists inhibit:Acute pain [Zimmer et al.]
Inflammatory pain [Clayton et al.]
Neuropathic pain [Ibrahim et al.]
• CB2 receptors located on:Immune cellsNeuronal cells (?) [Griffin et al; Ross et al; Patel et al.]
• CB2 agonists lack CNS side effects.
Development of potent selective CB2 ligands:Agonist: JWH-133 Ki 3.4nM with a 200-fold selectivity over CB1 receptors.Antagonist: SR144528 Ki 0.67nM with a 50-fold selectivity over CB1 receptors.
Aim: To determine the involvement of the CB2 receptor in nociceptive processing.
The Role of the CB2 Receptor in Nociceptive Processing:
An in vivo electrophysiological studyNeuronal Networks Laboratory
Combined unit / LFP with USV / behavioural recording
50 ms
25
50
75
100
125
kHz
1V
296:1
296
0.05 0.10 0.15 0.20 0.25 0.30 s
25
50
75
100
125
kHz
1V
295:1 295:2295:3 295:4 295:5 295:6
295
USV call start time
unit activity - nuc accumbens
USV Recorder input
Behavioural data
Local Field Potential (LFP)
Spectrogram of specific calls [AviSoft]
Behavioural Electrophysiology
Circular arena recording using CinePlex
Movie - rat HopScotch: 8-channel array in nuc. accumbens - 6 weeks post implant
Neuronal Networks Laboratory
Recorded video
Neural data
DISCUSSION • Following KA, hippocampal units (~80%) show an increase in firing; while mPFC units show either a decrease (~80%) or increase (~20%) in firing rate. mPFC units lose their characteristic bursting pattern after KA administration.
• CCH analysis shows that unit pair activity under basal conditions is more correlated within the mPFC compared to intra-hippocampal; mPFC appears to lead hippocampal firing. KA increased correlation within the hippocampus and mPFC; but the mPFC-hippocampal drive was lost.
• PDC of unit population activity also shows basal mPFC-hippocampal directionality (predominantly at low frequencies); this initially decreases after KA administration, then later increases at all frequencies.
• In basal conditions, PDC analysis of LFPs revealed evidence of information flow from CA3 to CA1 and reciprocal hippocampal-mPFC connectivity with predominant drive from mPFC to hippocampus. Following KA, there was increased drive from mPFC to hippocampus.
This alteration in functional connectivity in a seizure model has implications for memory and learning in epilepsy.
Caveat(s):
Need to consider possible influence of anaesthesia in directing “information flow” and/or (anaesthesia/KA-induced) short-term rewiring of neural circuitry. Other regions (not recorded) may be involved in communication between mPFC and hippocampus.