Han et al vs Yiu et al - Joe

Han et al vs Yiu et al

Seminar in Biopsych

Fall 2016

Rebecca Shansky

Diphtheria toxin receptors (DTRs) are not endogenously expressed in mice, and neither is diphtheria toxin (DT). Thus, inducing expression of DTRs in the LA in a cre-dependent manner allows for specific ablation of neurons in the LA through injection of DT anytime after ample infection of the virus. This is what Han et al did. Transgenic mice (iDTR mice) were injected stereotaxically with a cre-dependent virus. In the population of cells that the virus infected, cre was expressed, which means that DTR was also expressed. Thus, (intraperitoneal?) injection of DT anytime after the cre had had enough time to be expressed would induce apoptosis in the susceptible (DTR-positive) population. Up to this point, all the cells in the LA injection site expressed the DTR; however, if the cre virus also encoded cDNA for CREB, only the LA neurons that expressed high levels of CREB would also be infected with cre, thus inducing DTRs (introducing apoptosis susceptibility). This is interesting because they mentioned that the CREB-cre group and the control-cre group showed similar levels of cell death — if the CREB-cre group is supposed to infect a narrower subset of neurons and the same volume of virus is injected into both groups, one would imagine that DT injection would kill a smaller population of cells in the CREB-cre group.

It was convincing that there were several groups of controls in this paper. Firstly, there was the experimental group (CREB-cre with DT in iDTR mice), and the subsequent iDTR mouse controls (control-cre with DT and CREB-cre with vehicle). Another control could have been control-cre with vehicle, but if there was no experimental effect in the control-cre with DT group, the control-cre with vehicle probably would not have been very informative. Furthermore, there were wild type controls as a proof of principle that the CREB-cre virus was infecting cells that were preferentially active during fear conditioning (the cells that were encoding the memory). With all the controls, only the experimental group showed a significant change before and after DT administration. However, this entire paper is based on cell-type specific apoptosis and concurrent freezing behavior change, and no control was done to account for crude locomotor changes. The LA is just medial to the cortex, and CREB is expressed throughout the entire brain, and so ablation of cells expressing high levels of CREB can precipitate a number of confounds outside of the LA. This is without considering that probably not every cell in the LA that expresses high levels of CREB is recruited in a particular memory trace. Still, it controls for impairments in learning after ablation and impairments in other memories prior to ablation, which are very convincing. Still, this change in behavior could be due to a change in locomotor activity around ablation, which is accounted for by the brain days later.

An interesting question to ask is if the title of the paper suits the findings. The most convincing experiments in this paper were the controls for inabilities to learn after ablation and impairments to learn prior to ablation. This showed that if ablation is targeting a memory trace, it is not affecting other memories learned prior to the auditory fear learning, and it also did not actually damage the memory-making system in the brain for future use. Still, it is unclear whether the neuronal population annihilated specifically encoded that memory.

Yiu et al attempted to follow up the findings of Han et al and others to conclusively say how neurons in the LA are recruited to be a part of a particular memory trace. To test the claim that LA neurons are recruited based on intrinsic properties (their current excitability), Yiu et al employed similar methods to those that Han et al did; however, instead of completely eliminating a particular subset of neurons in the LA, Yiu et al modulated their neuronal excitability through various viral techniques. The best part about these first few experiments was that before they began to show how excitability ties into the recruitment of neurons for the memory trace, they did a series of proof of concept experiments showing that their methods indeed increase/decrease excitability reliably — good controls.

What was interesting about this paper was that the same claim was proven three times, using three different approaches; however, all three approaches told the same story: dnKCNQ infection increases neuronal excitability which preferentially recruits infected neurons into the memory trace; excitatory DREADD infection coupled with CNO administration increases neuronal excitability which preferentially recruits infected neurons into the memory trace; and lastly ChR2 infection with light administration increases neuronal excitability which preferentially recruits infected neurons into the memory trace. The only change between the experiments was the increase in temporal sensitivity of the experiments. Were they all necessary to drive that point home? I’m not sure. Nonetheless, the finding is awesome! Simply activating a subset of neurons before a learning task “primes them” and they subsequently are more likely to be a part of the memory trace, as measured by neuronal activity during a retrieval task (coupled with an in situ measuring arc translation). The experiments were well-controlled; they accounted for the active timeframe of HSV by doing behavior several days after injection of the virus, and also accounted for overall anxiety behavior by showing that increasing excitability in the LA doesn’t just alter locomotor behavior — it’s context specific, among several other crucial controls. Overall, it seems convincing that increasing neuronal excitability increases probability that the given neuron will become recruited into the memory trace. And it’s also crazy to get that insight into the underlying mechanisms of neuronal recruitment during learning.

Han et al had a question regarding what happens if the neurons that are recruited into a memory trace are eliminated from the mouse’s memory repertoire after fear conditioning; can they still recall the memory? Yiu et al took it a step back and said, “well what is it about these neurons that are recruited that allows them to be recruited; it could be any collection of neurons that becomes a memory trace, but if you induce excitability, can you artificially recruit those neurons in a memory trace?” Apparently, you can.