Abstract
Electrical and pharmacological stimulation methods are commonly used to study neuronal brain circuits in vivo, but are problematic, because electrical stimulation has limited specificity, while pharmacological activation has low temporal resolution. A recently developed alternative to these methods is the use of optogenetic techniques, based on the expression of light sensitive channel proteins in neurons. While optogenetics have been applied in in vitro preparations and in in vivo studies in rodents, their use to study brain function in nonhuman primates has been limited to the cerebral cortex. Here, we characterize the effects of channelrhodopsin-2 (ChR2) transfection in subcortical areas, i.e., the putamen, the external globus pallidus (GPe) and the ventrolateral thalamus (VL) of rhesus monkeys. Lentiviral vectors containing the ChR2 sequence under control of the elongation factor 1α promoter (pLenti-EF1α -hChR2(H134R)-eYFP-WPRE, titer 109 particles/ml) were deposited in GPe, putamen and VL. Four weeks later, a probe combining a conventional electrode and an optic fiber was introduced in the previously injected brain areas. We found light-evoked responses in 31.5% and 32.7% of all recorded neurons in the striatum and thalamus, respectively, but only in 2.5% of recorded GPe neurons. As expected, most responses were time-locked increases in firing, but decreases or mixed responses were also seen, presumably via ChR2-mediated activation of local inhibitory connections. Light and electron microscopic analyses revealed robust expression of ChR2 on the plasma membrane of cell somas, dendrites, spines and terminals in the striatum and VL. This study demonstrates that optogenetic experiments targeting the striatum and basal ganglia-related thalamic nuclei can be successfully achieved in monkeys. Our results indicate important differences of the type and magnitude of responses in each structure. Experimental conditions such as the vector used, the number and rate of injections, or the light stimulation conditions have to be optimized for each structure studied.
Highlights
Optogenetics have become a mainstream technique to modulate neuronal activity both in vivo and in vitro [1]
The motor circuit of the basal ganglia directs its output through the GPi and portions of the pars reticulata of the substantia nigra to the anterior portion of the ventrolateral thalamic nucleus (VLa) [10,11] Because of the reciprocal nature of these circuits and the complex trajectory of fibers of passage that pierce through the various basal ganglia nuclei, the interpretation of the results of conventional electrical stimulation methods is often complicated
Based on the pre-stimulation firing rate and coefficient of variation (CV, see Methods), striatal single unit recordings were divided into phasically active neurons (PANs) (n = 10), tonically active neurons (TANs) (n = 34), fast spiking neurons (FSNs), (n = 12) and unclassified (n = 36)
Summary
Optogenetics have become a mainstream technique to modulate neuronal activity both in vivo and in vitro [1] This technology, based on the delivery and expression of light-sensitive ion channel proteins (opsins) in specific neuronal populations, permits stimulation or inhibition of neurons with high temporal and spatial resolution [2,3]. We here report our experience with the use of optogenetic tools in the primate basal ganglia and related thalamic nuclei. These areas are involved in motor planning and execution, as well as non-motor functions, such as the learning of habits and procedures [6,7], and are the targets of many neurodegenerative disorders, including Parkinson’s and Huntington’s diseases [8]. The value of using the far more specific optogenetic techniques to study the physiology of the basal ganglia is well established in rodent studies [12,13,14,15,16,17]
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