Even though my summer research project can be summarized as simply running a seemingly endless amount of HPLC samples, such a description would not do service to the lab I’m a part of. What I will do is part of a larger project with a much wider scope. My summer project falls under the umbrella of a Tryptophan depletion study conducted in rat models which is a continuation of work done by my lab in the pas
Of particular interest in the etiology of depression and related affective disorders is the role of the serotonergic neurotransmitter system, the system responsible for the transport of serotonin/5-hydroxytryptamine (5-HT). Even though the exact underpinnings of serotonin’s role in mental disease are unknown, it has been confirmed to be involved with the onset of depression. The theory that depression involves a serotonin deficiency is still widely circulated but study results have sometimes produced contradictory and confounding results. Other neurotransmitter systems have been speculated in playing a role in affective disorder including the dopaminergic system. However, the serotonin deficiency theory is the most accepted and makes up the basis of Tryptophan depletion as a method of studying depression.
Due to its clinical significance, effective methods of studying depression in animal models have been the object of scrutiny in the past. One of the more relevant methods involves implementing a social defeat model: a small male mouse is put in an enclosure with a bigger male mouse which then proceeds to “beat up” the smaller mouse. This method leads to depressive behavior in the small mouse but such a model is not effective in female mice and thus excludes the gender that suffers the most from depression. To correct this problem, rapid tryptophan depletion (RTD) has been suggested as a method to induce depressive behavior in both male and female animals to study it.
Based on the serotonin deficiency theory of depression, RTD involves feeding animals (rats in our lab’s study) a mixture of several large neutral amino acids (LNAAs) which reduces the level of endogenous tryptophan (TRP) in the animal and subsequently the levels of 5-HT. TRP depletion relies on the fact that only a limited amount of LNAAs can pass through the blood brain barrier (BBB). The ingested LNAAs do two things to lower endogenous TRP and 5-HT levels: they stimulate protein synthesis and compete with endogenous TRP for entrance into the brain through the BBB. Because TRP is the amino acid precursor to 5-HT, reduced TRP levels theoretically lead to reduced 5-HT synthesis and depressive behavior.
My lab has done work with RTD in the past and right now they are focusing on studying possible differences of RTD between adolescents and adults. I have joined the lab just as they are in the middle of another RTD study and I will contribute the most in two different was: HPLC analysis and rat sacrifice and dissection. High performance liquid chromatography (HPLC) is an advanced analytical chemistry technique used to detect the presence of certain analytes in a sample. It relies on an electrode system to detect analytes as they pass through the column of the apparatus. Analytes take the form of different peaks in an HPLC program and the area is indicative of the concentration. My summer work deals mostly with the analysis of tryptophan, serotonin, dopamine, and the respective metabolites of these neurotransmitters.
Running samples through the HPLC apparatus makes of the bulk of my work but I also help in harvesting blood and brain samples from the rats used in the study. On my first day of brain dissection, my PhD supervisor even said I was a natural for rat brain dissection and that I knew how to chop a rat head just perfectly! What I think is hilarious, however, about analyzing rat brains using HPLC is that one must “homogenize” them. Rat brain matter on its own is too large for the apparatus so we liquefy them and essentially turn them into rat brain smoothies.
I hope to have analyzed enough samples to complete an interesting poster at the end of the summer. Even though this project has elements that might not align exactly with the research I want to do in the future, I hope to continue learning and enjoying this project like I already am.
