ASSEMBLY OF BRAIN CIRCUITS AND THE CELLULAR MECHANISMS OF BEHAVIOR
Our laboratory is interested in the assembly of neuronal circuits and the mechanisms by which brain circuits give rise to behavior. We focus on the process of neuron addition into the brain of vertebrates, and seek to understand how new neurons integrate into the circuits of the adult brain, and their role in information processing and storage. To address these questions our laboratory develops new technologies to genetically manipulate the development and biophysical properties of neurons. To investigate how behavior arises from the activity of neurons in brain circuits, we are generating transgenic songbirds to manipulate key genes involved in the assembly of circuits that mediate vocal learning behavior.
OVERALL RESEARCH SUMMARY
Most neurons in the brain are born before birth and are never replaced. In contrast, certain populations of neurons are continuously replaced throughout the life of the animal. Do neurons acquired in adult life participate in a special form of memory storage that requires the replacement of old neurons? In mammals, neuronal replacement occurs at high levels in two brain areas be involved in olfactory perception and spatial memory. In songbirds, the capacity to learn their songs varies during adult life, and this variation is correlated to radical structural changes in the brain nuclei controlling song, which include massive neuronal replacement. Recently we have developed several new tools that allow us to genetically control the function of neurons. By using these techniques we are manipulating the birth, death, and electrical function of newly generated neurons in the brain of behaving animals, both in the olfactory system of mice, and in the song system of songbirds.
Regulation of neuronal integration into brain circuits.
The brain of adult vertebrates harbors a population of neuronal stem cells that continues to proliferate throughout the life of the animal, and whose progeny migrate through the brain, differentiate into neurons, and establish synaptic contacts with other neurons in the circuit. We are interested in understanding the cellular and molecular mechanisms that control the integration of these neurons into neuronal circuits. We are currently testing the hypothesis that synaptic input into newly born adult neurons guides the integration of these cells into existing circuits. In addition, we are investigating the mechanisms that neurons use to adapt their intrinsic and synaptic properties as they integrate into circuits and communicate with other neurons. To study the role of electrical and synaptic activity on neuronal integration we have developed new tools to manipulate the biophysical properties of neurons by genetically modifying the activity of ion channels and neurotransmitter receptors.
Genetic control of the assembly of circuits involved in vocal learning.
Vocal learning depends on the ability of brain circuits to perceive and imitate sound sequences and use these sequences for communication. Songbirds such as canaries and zebra finches have been a favorite experimental system for the study of vocal learning in animals for decades. These animals exhibit a robust and spontaneous vocal learning behavior, and they have dedicated brain circuits, known as the song system, that participate in the learning and production of song. Zebra finches listen to the songs that their fathers produce, and imitate these sounds until they acquire a stable adult-like song. In this respect, the time course and strategy of vocal learning in zebra finches is very similar to the manner in which human infants learn to speak. These observations suggest that the zebra finch could be an ideal system where to start investigating the genetic and biological basis of vocal learning. Recently, our laboratory has succeeded in the development of a series of techniques that allow us to genetically modify the brain of songbirds. These technical advances open new opportunities for the study of the relationship between genes and learning in an animal species with a robust behavioral repertoire. We are currently generating transgenic songbirds to manipulate key genes involved in the assembly of circuits involved in vocal learning behavior.
Figure 1: Commitment of stem cells to the production of neurons with defined connectivity.
Stem cells located in the subventricular zone (SVZ) of adult mice are committed to generate the same neuronal type (granule cells in the olfactory bulb with defined connectivity. Stem cells located in the anterior regions of the SVZ (labeled in red) produce neurons whose dendrites reach the upper layers of the olfactory bulb. In contrast, stem cells located in the posterior SVZ (labeled in green) produce neurons whose dendrites branch in the lower layers of the OB. Our laboratory is investigating the molecules that regulate the patterns of connectivity of neurons in this brain region.
Figure 2: Regulation of synaptic connections by intrinsic electrical activity
Newly-generated neurons (green) in adult mice are rendered hyperexcitable by delivering into them Nachbac, a voltage-gated sodium channel, via recombinant retroviruses. (Bottom) Genetically-enhanced excitability increases the number of inhibitory synapses (arrows) on the genetically modified neurons (green). By genetically controlling the electrical properties of neurons we investigate how neuronal activity regulates the integration of cells into brain circuits, and the connections between neurons.
Figure 3: Subcellular distribution of synaptic inputs and integration of neurons into circuits (left) Granule cells in the olfactory bulb have neurotransmitter receptors in the apical (top) and basal (bottom) dendrites.
Recent evidence suggests that the inputs directed to the apical dendrites favors the survival and integration of new neurons into the bulb. In contrast, the input directed towards the basal dendrites impairs the integration of new neurons. We are currently investigating the molecular basis for this differential effect of these 2 subcellular compartments on the neuron’s survival.
Figure 4: Action potentials regulate the maturation of synaptic inputs.
We have recently generated reagents that allow us to genetically block action potentials in individual neurons. We have observed that neurons without action potentials (left bottom) survive and integrate into the brain, but they fail to receive synaptic input (right bottom). We are investigating the mechanisms by which intrinsic neuronal activity regulates the formation and functional maturation of synapses.
Figure 5: Genetically modified songbirds to investigate the molecular bases of vocal learning and complex behavior.
Our lab has developed several techniques to genetically manipulate the development and function of neurons during the assembly of neuronal circuits. We have recently developed new genetic methods that have allowed us to generate transgenic songbirds to investigate the genetic basis of the assembly of brain circuits involved in vocal communication. We are using these transgenic songbirds to investigate the rules by which neurons migrate, choose their final locations, establish connections with each other, and give rise to behavior.
Figure 6: Transgenic songbirds with a targeted mutation for the autism-related gene CNTNAP2.
We have generated transgenic songbirds targeting the autism-related gene CNTNAP2 and have observed that the mutant birds fail to accurately copy their tutor’s song. (Bottom right) Wild-type siblings copy the song of their fathers very accurately, but mutant birds fail to copy the parts of the song that are acoustically complex. We are investigating how perturbation of CNTNAP2 affects the assembly of brain circuits involved in learning and production of songs.
Figure 7: Genetic manipulation of activity and robustness of behavior.
We have observed that delivering a voltage-gated sodium channel to render neurons in the song system hyperexcitable leads to acute degradation of the song. Surprisingly, the song structure recovers very quickly (within 10 days) after the genetic manipulations. We are currently investigating the mechanisms responsible for this recovery.
Figure 8: A new genetic method to identify connectivity between neurons.
We have designed a new genetic strategy to identify the wiring diagram of brain circuits. The system is based on the logic of the delta-notch system and it will allows not only to trace connections between neurons, but also to genetically modify the physiological properties of circuits of connected neurons.