Understanding how different types of neurons function in networks and, ultimately, how they influence behaviour are key goals of neuroscience. It would be useful if the activity of particular types of neurons could be temporarily influenced in a reversible manner. Such a cell type-specific and reversible method for fine-scale analysis of circuitry, working on the fast signalling time scale of the brain, does not yet exist. Although genes can be turned on and off cell type-selectively, these methods applied to the mouse brain are too slow (days to weeks) to study fast-acting systems. Acute pharmacological intervention is the most promising strategy, because under acute blockade or stimulation there is little time for the system to adapt. Ideally, we would need fast acting drugs specific for selected cell types. The drug action should be immediate, reversible and should be terminated by giving an antagonist. Our strategy exploits the rich allosteric pharmacology of GABAA receptors to achieve selective and reversible attenuation and stimulation of mouse neurons by using genetically altered GABAA receptors activated by endogenous synaptically released GABA. The drugs zolpidem (Zo) and b-carbolines (bCs) enhance and inhibit, respectively, GABAA receptors containing a g2 subunit via the benzodiazepine binding site. We have made a mouse line insensitive to these drugs by changing the amino acid phenylalanine in position 77 (F77) to isoleucine (I) in the g2 subunit gene. Sensitiviy to Zo and bC can subsequently be restored to specific cell types in I77/I77 brains by expressing the wild-type g2 subunit with specific transgenes. The activity of the targeted cell types can be modulated by applying Zo or bCs. This method maintains all the advantages of a pharmacological approach (fast, reversible, and bi-directional drug action) combined with the specificity determined by the cell type selective expression of the drug target.
Engineering GABAA receptors to modulate the activity of selected types of neurons
OBERTO, Alessandra;
2004-01-01
Abstract
Understanding how different types of neurons function in networks and, ultimately, how they influence behaviour are key goals of neuroscience. It would be useful if the activity of particular types of neurons could be temporarily influenced in a reversible manner. Such a cell type-specific and reversible method for fine-scale analysis of circuitry, working on the fast signalling time scale of the brain, does not yet exist. Although genes can be turned on and off cell type-selectively, these methods applied to the mouse brain are too slow (days to weeks) to study fast-acting systems. Acute pharmacological intervention is the most promising strategy, because under acute blockade or stimulation there is little time for the system to adapt. Ideally, we would need fast acting drugs specific for selected cell types. The drug action should be immediate, reversible and should be terminated by giving an antagonist. Our strategy exploits the rich allosteric pharmacology of GABAA receptors to achieve selective and reversible attenuation and stimulation of mouse neurons by using genetically altered GABAA receptors activated by endogenous synaptically released GABA. The drugs zolpidem (Zo) and b-carbolines (bCs) enhance and inhibit, respectively, GABAA receptors containing a g2 subunit via the benzodiazepine binding site. We have made a mouse line insensitive to these drugs by changing the amino acid phenylalanine in position 77 (F77) to isoleucine (I) in the g2 subunit gene. Sensitiviy to Zo and bC can subsequently be restored to specific cell types in I77/I77 brains by expressing the wild-type g2 subunit with specific transgenes. The activity of the targeted cell types can be modulated by applying Zo or bCs. This method maintains all the advantages of a pharmacological approach (fast, reversible, and bi-directional drug action) combined with the specificity determined by the cell type selective expression of the drug target.
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