Injection of AAV8-GFAP-hM4Di-mCherry into the ACC resulted in expression of hM4Di (A) in 91.1 ± 2.4% of GFAP-positive cells (B) with 96.3 ± 0.9% specificity (C), whereas 3.6 ± 0.8% of NeuN-positive …
Zip file containing data for Figure 1B–D in GraphPad Prism file format.
(A) Representative sections 3.24 mm and 2.52 mm anterior to bregma showing hM4Di-mCherry expression (arrowheads) in the ACC. The arrow shows the site of microinjection and cannula placement into the …
(A) Performance index (PI) (mean ± SD) during the acquisition of the six original PAs (OPAs) (S1–2, 4–8, 10–17) and NPAs (S19) of the control (n=8), hM4Di-CNO (n=15), and rescue …
Zip file containing data for Figure 2A–O, Figure 2—figure supplement 2A–D, and Figure 2—figure supplement 3E–G in GraphPad Prism file format.
(A) Event arena for multiple flavor-place paired associate training. (B) Timeline for schema experiments.
(A) Performance index (PI) (mean ± SD) during the acquisition of the six original PAs (OPAs) (S1–2, 4–8, 10–17) and new PAs (NPAs) (S19) of the control (n=6) and control-CNO (n=4) groups. The …
(A) Timeline for the experiments. OFT was conducted for all groups (hM4Di-saline, hM4Di-CNO, and hM4Di-CNO+L-lactate; n=8 in each group) after two paired associate (PA) training sessions. …
(A) Experimental design to investigate the effect of Gi activation of ACC astrocytes on cAMP and L-lactate levels. (B and C) CNO decreases cAMP in the hM4Di-expressed cells. (B) Confocal micrograph …
Zip file containing data for Figure 3C–E in GraphPad Prism file format.
(A–C) Representative confocal micrograph of PGC-1α (A)/SIRT3 (B)/ATPB (C) co-labeled with NeuN, glial fibrillary acidic protein (GFAP), and DAPI (4′,6-diamidino-2-phenylindole) in the ACC of the …
Zip file containing data for Figure 4D–G in GraphPad Prism file format.
(A) Experimental design to investigate whether MCT2 and NMDAR activity are required for L-lactate-induced mitochondrial biogenesis. (B and C) mtDNA copy number abundance in the anterior cingulate …
Zip file containing data for Figure 5B–C and Figure 5—figure supplement 1B in GraphPad Prism file format.
(A) Western blot showing decreased expression of MCT2 in the anterior cingulate cortex (ACC) 12 hr after injection of MCT2 antisense oligodeoxynucleotide. (B) Intensity of MCT2 was quantified and …
L-lactate in the anterior cingulate cortex (ACC) is required for schema memory formation and neuronal mitochondrial biogenesis. Astrocytic Gi activation results in decreased L-lactate in the ACC …
Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
---|---|---|---|---|
Strain, strain background (Rattus norvegicus, male) | Adult Sprague- Dawley rats (250–300 g) | Laboratory Animal Services Centre, Chinese University of Hong Kong, SAR, China | ||
Transfected construct (Rattus norvegicus) | AAV8-GFAP-hM4Di-mCherry | Shanghai Taitool Bioscience Co. Ltd | ||
Transfected construct (Rattus norvegicus) | AAV8-GFAP-mCherry | Shanghai Taitool Bioscience Co. Ltd | ||
Sequence-based reagent | Rat MCT2 antisense oligodeoxynucleotide (ODN), 200 nmol (HPLC purified) | Integrated DNA Technologies (IDT) | Cat #: 107968967 | |
Sequence-based reagent | Rat Relative scrambled ODN, 200 nmol (HPLC purified) | Integrated DNA Technologies (IDT) | Cat #: 107969138 | |
Sequence-based reagent | Rat D-loop Forward and Reverse Primer | Integrated DNA Technologies (IDT) | Cat #: 107056074 and Cat #: 107056075 | Used to measure mtDNA by real-time PCR |
Sequence-based reagent | Rat β-actin Forward and Reverse Primer | Integrated DNA Technologies (IDT) | Cat #: 107056076 and Cat #: 107056077 | Used to measure nDNA by real-time PCR |
Antibody | Anti-GFAP (Mouse Monoclonal) | Abcam | Cat #: ab4648 | 1:500 (IHC) |
Antibody | Anti-NeuN (Rabbit Polyclonal) | Merck Millipore | Cat #: AB978 | 1:500 (IHC) |
Antibody | Anti-mCherry (Chicken Polyclonal) | Abcam | Cat #: ab205402 | 1:1000 (IHC) |
Antibody | Anti-cAMP (Rabbit Monoclonal) | Abcam | Cat #: ab134901 | 1:500 (IHC) |
Antibody | Anti-SIRT3 (Rabbit Polyclonal) | Sigma-Aldrich | Cat #: SAB5700222 | 1: 250 (IHC) |
Antibody | Anti-PGC-1α (Rabbit Polyclonal) | Abcam | Cat #: ab191838 | 1: 500 (IHC) |
Antibody | Anti-ATPB (Mouse Monoclonal) | Abcam | Cat #: ab14730 | 1: 500 (IHC) |
Antibody | Anti-MCT2 (Rabbit Polyclonal) | Merck Millipore | Cat #: AB3542 | 1: 500 (WB) |
Antibody | Anti-β-actin (Mouse Monoclonal) | Immunoway | Cat #: YM3028 | 1: 5000 (WB) |
Antibody | Alexa Flour 488 (Goat Anti-Mouse Polyclonal) | Thermo Fisher Scientific | Cat #: A11001 | 1:300 (IHC) |
Antibody | Alexa Flour 594 (Goat Anti-Mouse Polyclonal) | Thermo Fisher Scientific | Cat #: A11032 | 1:300 (IHC) |
Antibody | Alexa Flour 488 (Goat Anti-Rabbit Polyclonal) | Thermo Fisher Scientific | Cat #: A11034 | 1:300 (IHC) |
Antibody | Alexa Flour 594 (Goat Anti-Mouse Polyclonal) | Thermo Fisher Scientific | Cat #: A11037 | 1:300 (IHC) |
Antibody | Goat Anti-Rabbit Secondary Antibody, HRP, Polyclonal | Invitrogen | Cat #: 31460 | 1: 5000 (WB) |
Antibody | Goat Anti-Mouse Secondary Antibody, HRP, Polyclonal | Invitrogen | Cat #: 62–6520 | 1: 5000 (WB) |
Commercial assay or kit | Lactate Fluorescence Assay kit | Abcam | Cat #: ab65331 | |
Commercial assay or kit | cAMP complete ELISA kit | Abcam | Cat #: ab133051 | |
Commercial assay or kit | QIAamp DNA Mini Kits | QIAGEN | Cat #: 1725270 | |
Commercial assay or kit | SsoAdvanced Universal SYBR Green Supermix | Bio-Rad | Cat #: 1725270 | |
Commercial assay or kit | RIPA Buffer | Sigma-Aldrich | Cat #: 20-188 | |
Commercial assay or kit | Phosphatase and protease inhibitor cocktail | Sigma-Aldrich | ||
Commercial assay or kit | Bradford assay | Bio-Rad | Cat #: 5000205 | |
Commercial assay or kit | Western Bright ECL HRP substrate | Advansta | Cat #: K12045-D20 | |
Chemical compound, drug | Clozapine-N-oxide (CNO) dihydrochloride | Hello Bio | Cat #: HB6149 | |
Chemical compound, drug | NaCl | Sigma-Aldrich | Cat #: S3014-1kg | |
Chemical compound, drug | L-lactate | Sigma-Aldrich | Cat #: L-7022 | |
Chemical compound, drug | D-(-)-2-Amino-5-Phosphonopentanoic acid (D-APV) | Sigma-Aldrich | Cat #: A8054 | |
Chemical compound, drug | Artificial cerebrospinal fluid (ACSF) | Harvard Apparatus | Cat #: 597316 | |
Chemical compound, drug | Dorminal 20% | Alfasan International BV | Cat #: 013003 | |
Chemical compound, drug | Urethane | Sigma-Aldrich | Cat #: U2500-500G | |
Software, algorithm | FIJI ImageJ | National Institutes of Health, Bethesda, MD, USA | ||
Software, algorithm | Prism GraphPad | GraphPad Software, San Diego, CA, USA | Version 10 | |
Software, algorithm | Excel | Microsoft | ||
Other | Microdialysis guide cannula (CMA 11 elite) and probe (3 mm membrane) | CMA Inc | Used to collect ECF from ACC for L-lactate and cAMP assay | |
Other | Stainless steel guide cannulae, OD 0.41 mm-27G/C | RWD Life Science | Cat #: 62069 | Used in drug and ODN delivery into ACC |
Other | Dummy cannulae | RWD Life Science | Cat #: 62169 | Used in drug and ODN delivery into ACC |
Other | Brain slicer | Braintree Scientific, Braintree, MA, USA | Used to collect ACC from whole brain | |
Other | Stereotaxic frame | RWD | Used to fix head of rats during surgeries | |
Other | 33-Gauge metal needle, 10 μl micro-syringe | Hamilton, NV, USA | Used in AAV and drug delivery | |
Other | Microinjection pump | World Precision Instruments, USA | Used in AAV delivery |
Comparison of performance index of control vs. hM4Di-CNO group.
Comparison of performance index of control vs. rescue group.
Primer sequences and preparation of 20 μl reaction mixture for real-time PCR.