A Review on Optogenetics and Its Applications in Addiction Research
Writer: David James Estrin
Date: Fall 2016
Citation: Estrin, D. J. (2016). A Review on Optogenetics and Its Applications in Addiction Research. Rutgers Research Review, 1(2).
Optogenetics is a current tool used across disciplines to control membrane potential in genetically altered cells in vivo and in vitro. Dr. Gero Miesnböck introduced transgene expression in genetically modified mouse neurons to produce Channelrhodopsin-2 (ChR2) to control membrane potential via photostimulation, a procedure he later coined "optogenetics" (Zemelman et al., 2002; Lima & Miesenböck, 2005; Miesenböck & Kevrekidis, 2005; Miesenböck, 2009). By genetically modifying an animal's genome, scientists are able to photo-stimulate action potentials (APs) in distinct neurons subpopulations thus controlling the neuron's function.
To begin research using optogenetics, there are fundamental measures researchers take to construct discrete neurons whose APs are manipulated by light stimuli. By using the Cre-Lox recombinase method (Sauer, 1987), a targeted neuron's inserted ChR2 gene can be expressed in distinct neuron subpopulations possessing Cre recombinase (Tsai et al., 2009). To construct the Cre-responsive R26::ChR2-EGFP allele found on the GT(ROSA)26Sor (R26) locus, an inserted DNA fragment entailing 315 N-terminal amino acids produces a channelrhodopsin-2 protein when transcribed and translated (Boyden, 2011). Additionally, the DNA fragment inserted into the genome includes a ten-amino acid linker to an Enhanced Green Fluorescent Protein (EGFP) (Boyden, 2011). Finally, ChR2 is expressed when Cre-mediated excision of a loxP-STOP cassette interposes between transcription and translation start sites because a synthetic CAG promoter is activated (Boyden, 2011).
The primary use of inserting a ChR2-EGFP gene into a model organism's genome is to control action potentials (APs). APs are voltage sensitive and are created by minute changes in sodium, calcium and chloride concentrations on the axon membrane. ChR2 is a photostimulated voltage-gate which allows depolarization by calcium influx into a cell when stimulated. ChR2 has a similar mechanism as the G protein coupled receptor rhodopsin seen inside rod cells of the retina. By light exciting retinal molecules from the cis to trans isomer state, the ion channel opens allowing influx and efflux of ions (Figure 1, A-B). ChR2 is not the only photostimulated transmembrane protein gene utilized in optogenetics. Researchers commonly use archaerhodopsins and halorhodopsins to suppress neurons from firing action potentials (Boyden et al., 2005). With this in mind, by engineering a specific neuron to possess ChR2 proteins on the soma, axon hillock and unmyelinated axon, a neuron may be stimulated to fire an action potential by shining a high powered laser to activate ChR2 proteins (Kätzel et al., 2011). In summary, specific neurons like primary motor and somatosensory GABAergic interneurons may partially be controlled to fire by a laser stimulus (Kätzel et al., 2011).
Consequently, optogenetics is highly regarded in Neurophysiology and behavioral neuroscience research. Optogenetics is being used not only to understand the role of particular neural circuitry in the mammalian brain, but is a leading method in neural control prosthetics to combat disorders like addiction (Boyden et al., 2005). For instance, addiction to psychomotor stimulants such as cocaine and methamphetamine are understood to be controlled by the mesolimbic Pathway which is a projection from the ventral tegmental area to the nucleus accumbens. Another term for this reward circuit is the dopaminergic pathway because the neurotransmitter dopamine (DA) is commonly understood to control pleasure. When these narcotics are introduced, DA is unable to re-enter neuron or glial membranes leading to post-synaptic oversaturation of DA. This oversaturation will cause an individual to become inebriated. Because the mesolimbic pathway is strongly associated with addiction disorders, the National Institute of Drug Abuse is focused on using Optogenetics to both understand mesolimbic circuitry and ultimately suppress drug cravings in patients suffering chronic substance addiction disorders. By controlling firing patterns of specific mesolimbic neuron populations (e.g., medium spiny neurons or interneurons) that are associated with processing drug cues, one may hypothetically negate the affects of addiction. Thus, researches are aiming to suppress or enhance firing rates using optogenetics in specific areas of the striatum which may curb addiction.
Ultimately, the applications behind controlling action potentials by changing a neuron's genome have given researchers a new perspective in neuroscience research. By selecting specific light waves, rhodopsins and florescent proteins, scientists are beginning to further unravel the mysteries of the brain. As suggested, there is promise in using optogenetics to combat addiction in humans. Furthermore, optogenetics is being used to study many other disorders that affect the basal ganglia or the striatum.
Figure 1: (A) Rhodopsins are G protein coupled receptors which may be found in rod cell present on the retina. A Rhodopsin is comprised of the opsin protein and retinal molecule. (B) When retinal is stimulated by light it goes from a cis to trans formation. This change in stereochemistry open the ion channel. This change in ion concentration is sent from the retina towards the optic chiasma and brain. Figure adapted from Howgego & May, 2009.
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