Unraveling hippocampal networks related to food memories

第一个实验将是使用免疫组织化学来研究海马中Mc 4 rs和Leptin的重叠/非重叠表达。下丘脑中的两组神经元都编码饱腹感信号，并起到钝化进食的作用，当然它们具有不同的信号级联和电路机制。尽管这些受体都没有良好的抗体，并且鉴于低表达水平，原位几乎是不可能的，但我们可以在瘦素注射后将来自Mc 4 r启动子驱动的Cre系的GFP信号传导与STAT 3表达共定位（参见Myers等2012）。 接下来，显而易见的实验是用ChR 2或eNpHR来抑制Lepr+和/或Mc 4 r+海马神经元，并评估光刺激或光抑制对进食（量以及膳食模式，包括膳食大小、膳食频率、膳食长度）和体重的影响。如果我们看到一些令人信服的行为变化，那么我们就再次处于一个很好的位置，可以使用病毒工具来剖析电路的输入和输出。 更有趣的是执行条件位置偏好和其他记忆/觅食任务的能力，这些任务旨在在光遗传学操作期间获得食物。我们可以使用一个简单的腔室进行这些操作，甚至可以使用专门的Phenotyper笼设计自动化方案。 与上面列出的其他项目一样，我们可以（而且肯定应该）使用体外和离体切片记录的组合来分析这些细胞类型在不同状态（饥饿与饱足）或特定任务（记忆测定，主动进食等）中的放电模式。 工具 小鼠：Mc4r-t2a-Cre LepR-ires-Cre、Pacap-ires-Cre 病毒：AAV-FLEX-ChR 2-mCherry、AAV-FLEX-synapsin-mCherry、AAV-FLEX-eNpHR-YFP、AAV-FLEX-hM 3Dq-mCherry、AAV-FLEX-hM 4Di-mCherry、改良狂犬病病毒 设备：用于测量摄食量的表型分析笼。用于体内记录的Plexon Ominplex。急性切片ephys钻机。

The first experiment would be to use immunohistochemistry to investigate the overlapping/non-overlapping expression of Mc4rs and Leprs in the hippocampus. Both sets of neurons in the hypothalamus encode satiety signals and act to blunt feeding, of course with different signaling cascades and circuit mechanisms. Although neither of these receptors have good antibodies and in situs are nearly impossible given the low levels of expression, we can co-localized GFP-signaling from the Mc4r promoter-driven Cre line with STAT3 expression following leptin injection (see Myers et al 2012). Next, the obvious experiment is to transduce Lepr+ and/or Mc4r+ hippocampal neurons with ChR2 or eNpHR and assess the effects of photostimulation of photoinhibition on feeding (quantity as well as meal patterns including meal size, meal frequency, meal length) and body weight. If we see some convincing alteration in behavior, we are then in a great position again to dissect the circuit with regard to inputs and outputs using viral tools. Even more interesting is the ability to perform conditioned place preference and other memory/foraging tasks aimed at procuring food during optogenetic manipulation. We can perform these using a simple chamber or even design automated protocols using the specialized Phenotyper cages. As with the other projects listed above, we can (and definitely should) use a combination of in vitro and ex vivo slice recordings to analyze the firing patterns of these cell types in differential states (hungry versus sated) or specific tasks (memory assays, active eating ect). Tools Mice: Mc4r-t2a-Cre LepR-ires-Cre, Pacap-ires-Cre Virus: AAV-FLEX-ChR2-mCherry, AAV-FLEX-synapsin-mCherry, AAV-FLEX-eNpHR-YFP, AAV-FLEX-hM3Dq-mCherry, AAV-FLEX-hM4Di-mCherry, modified rabies virus Equipment: Phenotyper cage for measurements of food intake. Plexon Ominplex for in vivo recordings. Acute slice ephys rig.