Our objective is to use multidisciplinary approaches, from molecular biology, electrochemistry, genetics, pharmacology, to classical behavioural neuroscience techniques, to answer the questions that guide our research.
Below are synopses of techniques that we use in our laboratory. The methods we will employ will rarely be used in isolation; rather they will be used in conjunction with one another. It is likely that the list will be expanded to include other techniques.
Behavioural neuroscience begins and ends with a solid understanding of behavioural phenomena that give us insights into psychological and neural processes.
We use operant conditioning chambers equipped with auditory and visual stimuli and various manipulandae (e.g., levers, nose-pokes, wheels, and rods) on which rats can make responses to earn rewards, such as food treats or intravenous infusion of drugs (e.g., cocaine).
We also use mazes with sophisticated video tracking technology capable of triggering delivery of stimuli and rewards.
Optogenetics involves the generation of genetic constructs and insertion of these constructs into viral vectors (e.g., adeno-associated virus, herpes simplex virus, rabies virus). Subsequently, the viral vectors are infused into selective brain areas where, after a period of time, viruses induce the production and insertion of light-sensitive channels (opsins) into the neuronal membranes.
By shining laser light, through optical fibers positioned over neurons that express these opsins, we can control their activity (excitation or inhibition) with superior specificity and temporal resolution.
Chemogenetics – Designer Receptors Exclusively Activated by Designer Drugs (DREADDs)
Genetic constructs are created and inserted into viral vectors (e.g., adeno-associated virus). Subsequently, the viral vectors are infused into selectived brain areas where, after a period of time, viruses induce the production and insertion of novel neural membrane G-protein coupled receptors (GPCRs), such as hM3.
These receptors do not have endogenous ligands. Instead, the ligand for these receptors are designer drugs, such as Clozapine N-oxide (CNO). Importantly, CNO has no effect on organisms native receptors. By delivering CNO to select population of neurons expressing designer receptors, we can inhibit or activate them temporarily.
Fast-Scan Cyclic Voltammetry (FSCV)
FSCV is electrochemical detection technique used to measure neurotransmission with carbon-fiber electrodes. These microsensors, ~10 um in diameter and 130 um long, measure neurotransmitter release 10 times per second (i.e., 10 Hz).
By ramping up the electrical potential at the surface of the microsensor, catecholamines (e.g., dopamine) in the environment surrounding the microsensor oxidize. The ramp down reduces the analytes. This process elicits a small change in current which is measured by the electrode. Further, oxidation and reduction potentials are unique to specific analytes, which allows us to determine its identity.
Immunohistochemisty and Fluorescence Microscopy
Immunohistochemistry is a procedure of detecting and visualizing antigens in tissue. Specifically developed antibodies bind to antigens of choice and subsequently can be further tagged, with fluorescent or non-fluorescent tags, and observed under a microscope.
In cases of viral vectors (see above) staining of the tissue is not necessary as viral vectors induce the production of fluorescence (e.g., mCherry, EYFP).