pAAV-CaMKIIa-hChR2(H134R)-mCherry in Mouse (AAV9)
Addgene #26975-AAV9

In this image you can see the intense expression of the virus in Layers 4,5,6 of auditory cortex and hippocampus under green LED in 4X magnification. Despite the fact that under 4X we cannot see the virus expression, under 40X is apparent. See the next image....

Caption: In this image you can see the intense expression of the virus in Layers 4,5,6 of auditory cortex and hippocampus under green LED in 4X magnification. Despite the fact that under 4X we cannot see the virus expression, under 40X is apparent. See the next image....

Under 40X in Layer 2,3 of auditory cortex you can see the virus expression after photostimulating with green LED. Note that this image is coming from the previous one (at 4X magnification) in which the virus was undetectable in Layer 2,3)

Caption: Under 40X in Layer 2,3 of auditory cortex you can see the virus expression after photostimulating with green LED. Note that this image is coming from the previous one (at 4X magnification) in which the virus was undetectable in Layer 2,3)

In this image you can see an NMDA EPSC recorded from a SST interneuron (at +40 mV_VClamp) in Layer 2,3 of auditory cortex from the slice shown above. Note the small responses (around 40-50 pA) due to the insufficient virus expression.

Caption: In this image you can see an NMDA EPSC recorded from a SST interneuron (at +40 mV_VClamp) in Layer 2,3 of auditory cortex from the slice shown above. Note the small responses (around 40-50 pA) due to the insufficient virus expression.

Virus & Injection

Primary Virus
Promoter
CaMKIIa
Injected Titer
1.00E+13 GC/ml
Injected Volume
700 nl
Injection Rate
0. 1 μl/min
Injection Site
Brain parenchyma
Brain Region
Forebrain: Cerebral cortex (neocortex)
Injection Site Comments
I injected in auditory cortex targeting the excitatory axons of principal neurons (0. 5 to 1 mm deep from pial)
View in Addgene Catalog

Subject

Species / Strain
Mouse FVB-Tg(GadGFP)45704Swn/J obtained from Jackson Labs
Stock Number
3718
Age at Injection
21-26 days old

Data Collection

Time After Injection
3 weeks
Detection Method
Direct fluorescence, I recorded AMPA and NMDA EPSCs in SST and PV interneurons which were patched in voltage clamp.
Expression Site
Principal neurons in auditory cortex (axons mostly)
Genetic Tool Outcome
I ended up to inject to much volume of the virus from the original stock (without dilluting with 1XPBS) and to expect too many days (2 weeks and more) to get enough expression
Other Information
I did not see any toxicity but the virus expression was not enough
Tips & Comments
I would recommend to inject over 700 nl of the virus in the original stock (1E13) and to be very precise in the injection site. Obviously the virus does not spread a lot