CARTER, Adam, Synaptic Organization in Prefrontal Cortex, NYU, May 19, 2016
<p class="MsoNormal" >CARTER, Adam, Synaptic Organization in Prefrontal Cortex, NYU, May 19, 2016</p>
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4PM SNI talk at Stfd in S360...</p>
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rodent PFC is what he studies...</p>
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why is PFC important?</p>
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signal integration...</p>
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<b>PFC receives diverse excitatory inputs including cortex, HC= hippocampus, thalamus and BLA.
BLA = basolat. amygdala</b>.</p>
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top-down control..</p>
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PFC sends excitatory output to other areas.</p>
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so<b>, BIDIRECTIONAL communication</b>...</p>
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so, how do indiv neurons participate in long-range communic and circuits.</p>
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Methods</p>
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all in MICE... P28 to P56 (post-natal day)</p>
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in vivo injections</p>
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acute brain slices</p>
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whole cell recordings</p>
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2 photon microscopy and optogenetics</p>
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use fluorescent probes (imaged by 2PM)</p>
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identifying the unit...</p>
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Inject AAV-EGFP into one brain region and wait for expression and anterograde transport to PFC...</p>
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we can see axon profiles</p>
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PFC is agranular (NO layer 4) inputs come to all layers...</p>
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BLA inputs come into layer= L2 and also L1...</p>
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control inputs cPFC come into L1 and L2</p>
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and ventral HC inputs come into L5.</p>
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2nd anatomic feature...</p>
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we inject retrograde tags ... eg CTB cholera toxin subunits etc.</p>
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multiple projection neurons eg <b>CA cortico-amygdalar </b></p>
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<b>CS corticostriate</b></p>
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<b>CC = cortico-cortical</b></p>
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<b>CT = cortico-thalamic </b>in layer 5/6..</p>
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so reciprocal circuits in the PFC</p>
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today will talk about circuits involving</p>
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BLA
cPFC and
MD...=</p>
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<b>mediodorsal thalamus </b>, contralat PFC and basolat amygdala...</p>
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part 1</p>
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PFC amygdala circuits...</p>
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fear conditioning...</p>
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PFC communic bilat with BLA...</p>
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we focus on 2 cell types</p>
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<b>CA neurons = cortico-amyg and CC = cortico-cortical...</b></p>
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are these inputs biased towards or against projection neurons...</p>
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se how to ID these?</p>
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we inject green retrobeads in BLA or cPFC = contralat PFC.</p>
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CA and CC neurons have indistinguishable morpology...</p>
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exactly the same... dendrites go up into layer 1/2</p>
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and have 6 mm of dendrites...</p>
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we patch them...</p>
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same morphology and same physio...</p>
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but how are they contacted by BLA = basolat amyd.</p>
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we look at synaptic connex...</p>
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look at neighboring neurons to compare (to eliminate the confounds)</p>
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directly compare synaptic responses ,,
excit inputs from CA neurons (compared to CC contacts)</p>
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only for the amygdalar inputs...</p>
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so, why are there larger inputs from amydala..</p>
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<b>synaptic strength depends </b>on</p>
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<b>1) prob of release</b></p>
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and <b>2) unitary response size </b>A</p>
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and <b># of connex N</b></p>
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EPSC = <b>product of the 3 = NQPr</b>....</p>
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so, 1) how abt presynap release prob Pr...</p>
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but presyanp excit from ChR is different than physio activation!</p>
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usually exaclty the same btwn CC and CA (so THAT's not the answer)</p>
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JH: John Huguenard... A: ignore stim duration and EPSC amplitude...</p>
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we treat the BLA inputs as a homogeneous group (and ignore diffs)</p>
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</p>
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how abt Q the post synap factors</p>
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we include strontium (similar to Cal) but it stays in.</p>
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compare quantal release...</p>
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yes. quantal release is bigger for CA... so that's a contrib factor but NOT the whole story</p>
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how about...</p>
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*</p>
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3) the # of inputs... (work of Justin Little)</p>
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map synaptic connex across the dendrites</p>
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use 2P imaging to</p>
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see post synaptic calcium signal...</p>
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flash blue light onto a section of dendrite...</p>
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and see spines...</p>
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he shows images of spines on dendrites..</p>
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flash blue light
and see spines lighting up...</p>
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those ARE activated by us...</p>
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justin did these for pairs of CA and CC inputs...</p>
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<b>there are MORE synapses on the CA neurons</b>!</p>
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and spine volume is large...</p>
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so, <b>differences in synaptic density and sub-cell targeting is the answer</b>...</p>
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shows data on connectivity (N)</p>
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<b>yes... spine density is the big difference</b>.</p>
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finally...</p>
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do they differentially drive these neurons</p>
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we can never fire CC neurons but <b>we CAN fire CA neurons</b>...</p>
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part 1 conclusion</p>
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CA and CC neurons are distinct cell types (despite having the same morphology)</p>
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BLA inputs are biased onto CA neurons</p>
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BLA inputs do self drive CA neurons.</p>
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(so, sort of dangerous reciprocal bidir communic btwn pfc and amyd)</p>
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BUT BLA seems to gen a large inhib response in PFC...
*</p>
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part 2.. inhib circuits in the PFC...</p>
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but many difft cell types in PFC</p>
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eg CA, PV, SOM+, CC</p>
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so how do BLA inputs evoke inhib in PFC?</p>
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all of our data says NON-volume xmision... rather it is specific point to point xmission.</p>
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now we focus on infralimbic PFC...</p>
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bulk of BLA input goes to infralimbic...</p>
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so these are compared to CS = cortico-striatal neurons...</p>
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so, what does activation look like</p>
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hold at -70 mv with patch...</p>
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we still have excit in CA... and lag of 2 msec...</p>
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but inhib is also biased.</p>
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both excit and inhibition appear stronger at CA neurons...</p>
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are inhib inputs biased toward CA neurons?</p>
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use transgen lines..</p>
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use pV CRE lines and SOM+ cre line.. interneurons...</p>
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we always use AAV injecs...</p>
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PV and SOM+ interneurons are similar to other in cortex...</p>
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studying inhib connex.</p>
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we do dual retrograde</p>
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AAV Ch@ into PFC.. flash light</p>
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PV and som+ both...</p>
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much larger onto the CA neurons than onto the CS neurons.</p>
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shows IPSCs at CA and CS neruons...</p>
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yes... inhib is also biased onto CA interneurons...</p>
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did conditional rabies...</p>
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use TVA=cherry and FLEX rg and after a week.. another agen..</p>
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both PV and SOM get inputs fro BLA...</p>
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recorded at -70mV</p>
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for CA and at ... to record PV+ neruons...</p>
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BLA inputs make stronger synapses onto PV interneurons</p>
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inputs from BLA depress both the PV and the SOM interneurons in PFC</p>
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remarkably similar...</p>
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compares AMPA dn NMDA response...</p>
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so BLA input is similar at CA neurons for both...</p>
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presyanp properties...</p>
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they depress the CA neurons but they facilitate the SOM+ interneurons...</p>
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yes presyanp properties do differ.</p>
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BLA inpts ot PFC so really big for PV+ interneurons...</p>
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and temporal sequence?</p>
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*</p>
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look at all 4 difft types of neurons</p>
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CS, CA, PV and SOM...</p>
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which fires first?</p>
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CA, PV and SOM all fire but we can never get the CS neurons to fire.</p>
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order...</p>
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PV neurons fire first and then SOM and then the CA neurons (in terms of current intensity)</p>
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we do not allow polysynaptic activity...</p>
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PV neurons always fire first.. rapidly makes cells fire...</p>
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SOM interneurons also fire well before CA neurons...</p>
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so... SLA inputs ot PFC</p>
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first PV then SOM and then CA (but never CS)</p>
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BLA inputs is primarily INHIBITING the PFC... so, why isn't it just blocked?</p>
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how abt VIP neurons... (another type of inhib interneuron)</p>
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no... nothing in VIP interneurons...</p>
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part 2 conclusions...</p>
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yes.. BLA strongly activates PV and SOM interneurons</p>
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and BLA does not engage a disinhibitory netowrk</p>
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(interneurons are also biased onto CA neurons)</p>
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*</p>
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part 3...</p>
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cortical and thalamic circuits...</p>
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do the above also apply to...</p>
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PFC-thalamus (recipr)</p>
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and also CC (cortico-cortical recip circuits)</p>
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we use all our tools look at these also...</p>
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compare CC and CT...</p>
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CT inputs classical layer 5 ...
and massive apical dendrite.</p>
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CT neurons...</p>
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CC and CT neurons have distinct morpho and physio.</p>
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these 2 neighbors look and act v different.</p>
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*</p>
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speed diffs and threshold and resp diffs.</p>
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thalamic inputs are larger the further out you go onto the dendrite for the CT inputs...</p>
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*</p>
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cortical inputs...</p>
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actual EPSC for both CC and CT actually seem to be the same</p>
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despite big diff in input resistance...</p>
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part 3 conclusions...</p>
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CC and CT neurons are distinct cell types...</p>
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EPSPs are enhanced at CC neurons but faster at CT neurons.</p>
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*</p>
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future...</p>
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10 difft pyrams in layer 5.</p>
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who connects onto them?</p>
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2) how are these circuits modulated? (acute and chronic eg dopamine modulate...</p>
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D1+ neurons... </p>
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but D1+ and D1- neurons are v distinct morpho and phys...</p>
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finally how does PFC influence downstream targets..</p>
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in striatum, amyg and thalamus?</p>
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shows lab members...</p>
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Q abt diff pops in BLA..</p>
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but always get recip back from PFC.</p>
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Q subcell cell bio...</p>
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how do they engage difft dendriticproperties..</p>
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PFC is highly recurrent... so hard to activate input without activ the recipr connex.</p>
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hard to look at function of thalam cortical... even if u crnak thelight way up..</p>
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he gave up trying to use spectrally distinct light... too difficult (he's too old andjaded)</p>
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we think PV+ and SOM+ are distinct genetically.</p>
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CAM may dictate the difference.</p>
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input from BLA to layer 2... important</p>
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(how abt layer 5 inputs from BLA asks JH)</p>
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A not explored...</p>
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Q how abt VIP neurons.. why no response?</p>
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A perhaps its another input that can engage the disinhib input...</p>
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