journal club #5 - in vivo inhibitory activity
DESCRIPTION
In Journal Club #5, Mac Hooks takes us through some recent results from Carl Petersen's lab, on the membrane potential dynamics of GABAergic neurons in the barrel cortex of behaving mice.TRANSCRIPT
Membrane potential dynamics of GABAergic neurons in barrel cortex of behaving mice
Gentet et al. (Petersen lab) Neuron 65: 422 (2010)
Mac HooksJournal Club #5
MethodsGABAergic input suppresses AP firing in L2/3 pyramids
All whiskers except C2 trimmed
Awake head restrained GAD67-GFP knockin mouse; habituated to restraint
Location of C2 barrel via intrinsic optical imaging
In vivo 2p imaging of GFP+ GABAergic neurons
Whole cell recording (K-gluconate; Alexa 594; biocytin)
Recordings L2/3 (about 180-220µm deep)
Sac after recording for anatomy
GABAergic input suppresses AP firing in L2/3 pyramids
Slow, large amplitude Vm fluctuations
Note AP frequency shiftQuantification of AP frequency shift
Mode shift to “bursting” (2 or more APs) in gabazine
“Quiet wakefulness”
This shows that GABAergic transmission is capable of modulating firing, but not which circuits are engaged (note later that AP firing in excitatory is not affected by shift from quiet to active wakefulness)
GAD67-GFP knockin mouse Tamamaki et al. J Comp Neurol (2003) 467:60
Two lines:GAD67-GFPGAD67-GFP(Δneo): mated to CAG Cre mouse to eliminate loxP-flanked PGK-neo cassette (in case it affects expression)
(Almost) all GFP+ neurons are GABAergicFigure S1 from Petersen (left)
Tamamaki (below ,neocortex) )gives GFP+ with GAD67 (156/196) and no GFP+/GAD- neurons
GAD67-GFP knockin mouse:Interneuron Subtypes (Motor cortex)
Characterization of Interneuron Subtypes in Frontal/Motor Cortex
All GFP+ cells are NeuN+ (about 19.5% of cortical neurons)
L1/2
L2/3
L2/3
L5/6
GAD67-GFP knockin mouse:Interneuron Subtypes (S1 cortex)
Figure S5
Tamamaki et al.
marked for comparison
Whole-cell recordings of GABAergic neuronsClassifying 3 cell types
GFP+ GFP+
Ok. Should we ask for a more fine division of interneuron population?
AP half width AP frequencyResting VmInput resistance
Classifying 3 cell types, Quantified
Whole-cell recordings of GABAergic neurons
Fig S3 also adds more data ….
Behavioral modulation of Vm dynamics in GABAergic neurons
Examples
Tracking whisker angle (video; 20s limit)
Vm (aligned)Expanded to show subthreshold
Quantify:AP frequency
Vm
Variance of Vm
1-5Hz area (V dot Hz) of Vm fourier transform
Divide time into (Q) and (W)
Behavioral modulation of Vm dynamics in GABAergic neurons
3 cell types, quantified
Behavioral modulation of Vm dynamics in GABAergic neurons
Fast (subthreshold) membrane potential oscillations phase locked to whisking
Whisker angle
Average Vm(aligned to peak of protraction)
Peak amplitude of fast (subthreshold) membrane potential oscillations, plotted at the time in the whisking cycle where it
occursNO DIFFERENCES BETWEEN CELL
TYPES
Correlated activity of excitatory and inhibitory neurons during (Q) quiet wakefulness
Two excitatory neurons(Dual in vivo patch: distance averaged 140±19 μm; in a 300 μm diameter barrel, this could put one in
the middle and the other at the edge)
Cross correlogram
Essentially examining subthreshold correlation
Does whisker trimming affect correlation across cortex?
Correlated activity of excitatory and inhibitory neurons during (Q) quiet wakefulness
Highly synchronous slow oscillations
Interneurons then can’t drive the hyperpolarized
phase in pyramids …
Behavioral modulation of correlated activity
Behavioral modulation of correlated activity
Why not subdivided for FS/non-FS?
Large brief specific events
Action potentials in excitatory neurons are driven by large, brief, cell-specific depolarization
(e.g. events in excitatory neurons are not correlated)
Shuffled events
Histogram of relative spike times
(10ms bins)
Data
Essentially examining suprathreshold correlation
Action potentials in inhibitory neurons are driven by broadly synchronized depolarization
Synchronized depolarizations
Summary of Vm prior to APs
Spike triggered averages for …
Spikes triggered in excitatory neurons include events not present in …
…excitatory, or… FS interneuron
Whereas these events are more similar (e.g. broad depolarizations)
Vm slope in 20ms prior to spike initiation
Is there any way this could be some artifact of the cell type?
A Model for Behavioral Modulation of Vm Dynamics
But …
(3) L3→L3 connections are among the strongest excitatory local
connections … wouldn’t we expect the most synchrony in excitatory
neurons of L3 and L4? Or does the sparseness of connectivity explain?
(1) Is there any evidence for the shift in excitatory input from FS to Non-FS interneurons during activity?
Could we look for this in pathways (POm?) that might be more excited during active whisking?
(2) How does this model account for the shared broad depolarizations of FS and Non-FS interneurons? (is the “shared input” from L3 pyramids or elsewhere)
Feedforward inhibition engaged by thalamocortical inputCruikshank et al. (2010) Neuron 65:230
Connors’ data for VB feedforward to L4 is similar to what Petersen proposes for L3:
-general principle of feedforward from layer to layer?
-would POm input be more similar to ‘active’ wakefulness, while VB is more similar to ‘quiet’
wakefulness?