Also in the Article



Experiments were performed on brain slices from 12- to 15-week-old genetically marked (with ZsGreen) Glp1r- and Lepr-expressing POMC neurons using POMCDre LeprCre ROSA26lSlrSrZsGreen+/− or POMCDre Glp1rCre ROSA26lSlrSrZsGreen+/− male and female mice. Animals were kept under standard laboratory conditions, with tap water and chow available ad libitum, on a 12 h light/dark cycle. The animals were lightly anesthetized with isoflurane (B506; AbbVie) and decapitated. Coronal slices (270–300 µm) containing the ARC were cut with a vibration microtome (HM-650 V; Thermo Scientific) under cold (4 °C), carbogenated (95% O2 and 5% CO2), glycerol-based modified artificial cerebrospinal fluid (GaCSF)50. GaCSF contained (in mM): 244 glycerol, 2.5 KCl, 2 MgCl2, 2 CaCl2, 1.2 NaH2PO4, 10 HEPES, 21 NaHCO3 and 5 glucose, adjusted to pH 7.2 with NaOH. If not mentioned otherwise, the brain slices were continuously superfused with carbogenated aCSF at a flow rate of ~2.5 ml min−1. aCSF contained (in mM): 125 NaCl, 2.5 KCl, 2 MgCl2, 2 CaCl2, 1.2 NaH2PO4, 21 NaHCO3, 10 HEPES and 5 glucose, adjusted to pH 7.2 with NaOH. To block GABAergic and glutamatergic synaptic input, in all recordings, the aCSF contained 10−4 M picrotoxin (P1675; Sigma-Aldrich), 5 × 10−6 M CGP (CGP-54626 hydrochloride; BN0597, Biotrend), 5 × 10−5 M DL-AP5 (DL-2-amino-5-phosphonopentanoic acid; BN0086, Biotrend) and 10−5 M CNQX (6-cyano-7-nitroquinoxaline-2,3-dione; C127, Sigma-Aldrich). To suppress action-potential-dependent synaptic release, blocked voltage-dependent Na+ channels were blocked by 10−6 M TTX (T-550, Alomone).

Note: The content above has been extracted from a research article, so it may not display correctly.



Also in the Article

Q&A
Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.



We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.