PMID- 22866030
OWN - NLM
STAT- PubMed-not-MEDLINE
DCOM- 20121002
LR  - 20211021
IS  - 1662-5110 (Electronic)
IS  - 1662-5110 (Linking)
VI  - 6
DP  - 2012
TI  - An amplified promoter system for targeted expression of calcium indicator 
      proteins in the cerebellar cortex.
PG  - 49
LID - 10.3389/fncir.2012.00049 [doi]
LID - 49
AB  - Recording of identified neuronal network activity using genetically encoded 
      calcium indicators (GECIs) requires labeling that is cell type-specific and 
      bright enough for the detection of functional signals. However, specificity and 
      strong expression are often not achievable using the same promoter. Here we 
      present a combinatorial approach for targeted expression and single-cell-level 
      quantification in which a weak promoter is used to drive trans-amplification 
      under a strong general promoter. We demonstrated this approach using recombinant 
      adeno-associated viruses (rAAVs) to deliver the sequence of the GECI D3cpv in the 
      mouse cerebellar cortex. Direct expression under the human synapsin promoter 
      (hSYN) led to high levels of expression (50-100 muM) in five interneuron types of 
      the cerebellar cortex but not in Purkinje cells (PCs) (</=10 muM), yielding 
      sufficient contrast to allow functional signals to be recorded from somata and 
      processes in awake animals using two-photon microscopy. When the hSYN promoter 
      was used to drive expression of the tetracycline transactivator (tTA), a second 
      rAAV containing the bidirectional TET promoter (P(tet)bi) could drive strong 
      D3cpv expression in PCs (10-300 muM), enough to allow reliable complex spike 
      detection in the dendritic arbor. An amplified approach should be of use in 
      monitoring neural processing in selected cell types and boosting expression of 
      optogenetic probes. Additionally, we overcome cell toxicity associated with rAAV 
      injection and/or local GECI overexpression by combining the virus injection with 
      systemic pre-injection of hyperosmotic D-mannitol, and by this double the time 
      window for functional imaging.
FAU - Kuhn, Bernd
AU  - Kuhn B
AD  - Princeton Neuroscience Institute, Princeton University Princeton, NJ, USA.
FAU - Ozden, Ilker
AU  - Ozden I
FAU - Lampi, Yulia
AU  - Lampi Y
FAU - Hasan, Mazahir T
AU  - Hasan MT
FAU - Wang, Samuel S-H
AU  - Wang SS
LA  - eng
GR  - R01 NS045193/NS/NINDS NIH HHS/United States
GR  - RC1 NS068414/NS/NINDS NIH HHS/United States
PT  - Journal Article
DEP - 20120731
PL  - Switzerland
TA  - Front Neural Circuits
JT  - Frontiers in neural circuits
JID - 101477940
PMC - PMC3408591
OTO - NOTNLM
OT  - AAV
OT  - FCIP
OT  - GECI
OT  - TET
OT  - cerebellum
OT  - crus
OT  - in vivo
OT  - two-photon
EDAT- 2012/08/07 06:00
MHDA- 2012/08/07 06:01
PMCR- 2012/01/01
CRDT- 2012/08/07 06:00
PHST- 2012/03/10 00:00 [received]
PHST- 2012/07/09 00:00 [accepted]
PHST- 2012/08/07 06:00 [entrez]
PHST- 2012/08/07 06:00 [pubmed]
PHST- 2012/08/07 06:01 [medline]
PHST- 2012/01/01 00:00 [pmc-release]
AID - 10.3389/fncir.2012.00049 [doi]
PST - epublish
SO  - Front Neural Circuits. 2012 Jul 31;6:49. doi: 10.3389/fncir.2012.00049. 
      eCollection 2012.