Guest Post: A Year in Neuroscience

Studying the mammalian brain system was never part of my so-called "academic plan." It just happened to be a better option than being a newcomer in the job market — for me at least. So, I ended up in a Brain Research Institute neuroscience lab for a year, with an interesting research topic entitled, "Expression of clock genes in the brain during depression." The project revolved mainly around the effects of depression on the human clock, which many scientists believe lies in a brain area called the Suprachiasmatic nucleus (SCN). This tiny segment of the brain (part of the hypothalamus) is essential. It regulates our daily routines such as sleeping, eating, and other various cycles of the body. I like to think of the SCN as kind of orchestra conductor, as it delicately serves its function by coordinating different organs of the body.

Each side of the SCN consists of approximately 10,000 neurons and expresses clock proteins involved in the regulation of the mammalian circadian system. It can work both directly or indirectly through damped/peripheral oscillators. This may sound complicated, but the oscillators can simply be regarded as small units similar to the SCN, which take their orders from the latter to keep the body in phase with its 24-hour cycle.

For my part, I was mostly interested in the resulting effect of depression on the expression of clock genes in the 24-hour body cycle. The cool thing was that the project would include lab work involving both behavioral tests and molecular work. My experimental subjects were rodents (C57BL/6N male mice). Do mice get depressed? Well, it turns out you can simulate the brain chemistry of depression in mice by treating young mice with Citalopram, a selective serotonin reuptake inhibitor (SSRI). Some amazing research performed with SSRIs has shown that if Citolapram is given acutely during the postnatal phases, mice are more likely to be depressed when they become adults. Their behaviors are also subsequently altered. For instance, these mice can develop a high level of anxiety. This effect may be due to the fact that treatment with an SSRI during development can alter the serotonergic system. What does this have to do with humans? While doing some background reading, I came across some papers that mentioned increased teenage suicidal tendencies in teenagers who were exposed to antidepressants during prenatal development (their mothers took SSRIs during late pregnancy) — food for thought for future mothers.

The first battery of tests I conducted was behavioral. One was an open-field test (OFT) and the other was daily activity monitoring. My aim was to determine whether the Citalopram-treated mice were indeed expressing depression-like symptoms or any other anomalies in the daily routine. OFT was uber cool. Rodents that display signs of anxiety show alternate pattern of locomotion (quite simply, they move differently) as compared to normal (control) animals, due to their reduced exploratory behavior. Indeed, they tend to prefer peripheral movement (see image). Daily activity monitoring meant I drew a 24-hour activity profile of the rodents' activity, which ultimately measured their morningness and eveningness. We later compared our observations from the Citolapram-treated and normal groups to see if there were any differences. If the SSRI-treated mice were different from controls, this could reflect a change in the expression of clock genes. From these behavioral experimental results, we found that the acute exposure to SSRIs during the postnatal stage did have repercussions in the adult mice. We observed symptoms such as anxiety and circadian irregularities, while sexual dysfunction and depression-like behaviors were not definitively obvious. To definitively determine any changes in those latter behaviors, we suggested that further behavioral examinations were needed.

After the behavioral experiments, we did some molecular analyses, such as assessment of gene expression levels using real-time PCR. Unfortunately, after two months of the behavioral observations, the mice had to be sacrificed so we could do the molecular analysis of their brain tissue. But the exciting results overwhelmed my initial sadness. Upon investigating the expression of clock genes in brain regions such as the preoptic-area, we discovered that there was no significant change in expression level in both treated and control animals. The upshot was that we saw no changes in gene expression in the brain areas we analyzed.

What's the connection between the altered locomotion pattern, the circadian components, and postnatal antidepressant treatment? No one is sure, but one hypothesis about the chemical mechanism linking these concepts would be an increase of serotonin levels within the synaptic cleft during early-life SSRI exposure could in turn increase activation of post-synaptic serotonin receptors. This effect could cause the malfunction in the clock circuitry and be further expressed by the down-regulation of expression of certain genes dependent on this circuitry, such as arginine vasopressin (AVP). This particular protein might be involved in locomotion, which might account for the behavioral changes seen in the Citalopram-treated male mice. However, determining a definitive conclusion mechanism would require further experimental work. So this is the best conclusion we can make so far: Postnatal exposure to Citalopram does affect the behavioral components of the mice. How involved the clock genes are in this phenomenon is however still unclear.

That one year of assiduous research was the best experience I could have possibly obtained in my young academic career. Besides the guidance from experts in the related field, I could feel that everything was at the tip of my fingers: knowledge about things unknown, cutting-edge technology. I also realized that the constant support of friends and close ones was crucial in the sometimes frustrating field of neuroscience research (and I would anticipate scientific research itself).

Neuroscience is similar to an intoxicating substance — once you are in it, there's no chance of backing off! That's why my so-called academic plan shall be further modified to maybe fit it some more years of neuroscience research.

--Manish Putteeraj

Image Credit: Dr. Tomoko Soga (Brain Research Institute, Monash University)