Garner Lab Research Areas
Human neural development is an amazingly complex process. It begins with the birth of neural cells in the developing neural tube and is followed by the migration of these cells to specific layers within the developing brain. Near the end of their migration nerve cells undergo a phase of differentiation during which they form both axons and dendrites that traverse large distances forming synaptic contacts with specific subpopulations of target nerve cells throughout the central and peripheral nervous system. (EM Picture of a synapse). Mechanistically, these early phases of neuronal differentiation and synaptogenesis appear to be largely genetically encoded, however once axons reach their appropriate targets, the refinement, maintenance and elimination of synaptic contacts becomes strongly influenced by neural activity and sensory input. These activity dependent events are tightly coupled to a phenomenon, known as synaptic plasticity, a process wherein synapses can modify the strength with which they transmit information. For example, strong correlated signaling tends to strengthen synaptic contacts, while weak asynchronous neurotransmission lead to the weakening and/or loss of synapses. This process of synaptic plasticity is also critically important for the encoding and processing of information in our mature brain and is ultimately responsible for cognition and our ability to learn, retain and access memories.
Of the many phases of neural development, we are most interested in understanding the molecular mechanisms underlying the formation and plasticity of synapses, as these events are critical to the proper wiring of the nervous system and for information processing in the developing and mature brain. In particular, we are and have been interested in addressing a number of fundamental questions. First, what molecular mechanisms guide the assembly of synapse? Second, how do changes in neural activity modulate synaptic strength? Finally, how do genetic or environmental insults to the developing nervous system result in impairment in cognitive function. Intriguingly, experiments designed to address each of these question have begun to lay the foundation for both an understanding of how specific genetic lesions can cause intellectual disabilities and possible treatment strategies.
Functional assembly of CNS Synapses
Achieving a fundamental understanding of how cognition is encoded by the brain requires insights into the molecules that define synaptic junctions and their roles in regulating neurotransmission. As such we have devoted significant effort during the last two decades in identifying and characterizing proteins of vertebrate CNS synapses. Initial clues to the molecules that comprise synapses of different types came in the late eighties through a fantastic collaboration with Dr. Eckart Gundelfinger currently a Professor at the Leibniz Institute for Neurobiology in Magdeburg Germany. Here our two groups searched for the first synaptic junctional proteins by screening cDNA expression libraries with polycolonal antibodies against rat brain synaptic junction in collaboration with Eckart Gundelfinger. This led to the cloning of cDNAs to more than 80 novel synaptic proteins. Since that time, we have worked together to evaluate the functional importance of many these pre and postsynaptic including Piccolo, Bassoon, SAP90, SAP97, SAP102, ProSAP.
Using a combination of biochemical, cellular, molecular, electrophysiological and reverse genetic approaches we have found that each of these are scaffold proteins of primarily glutamatergic synapse and play fundamental roles in the nascent assembly and function of these synapse. For example, members of the SAP90/SAP97 and SAP102 family of MAGUKs have been found to function primarily postsynaptically in the recruitment, localization and retention of glutamate receptors. In contrast, the presynaptic active zone proteins Piccolo and Bassoon have been found to facilitate the assembly of the active zone and to regulate the release of neurotransmitters.
Synaptic protein dynamics
As our studies of individual synaptic proteins progress, we became increasingly interested in questions related to the dynamics of synapse formation. For example, how long does it take a nascent synapse to form? How are pre and postsynaptic proteins delivered to nascent synapses? Once a synapse is formed, do the proteins that comprise the junction exchange and on what time scale: seconds, minutes, hours, days months or years? How do these exchanges kinetics relate to the life-time of individual synapses? What mechanisms regulate synapse stability and elimination?
To address these and related questions, we embarked on a second fantastic collaboration with Dr. Noam Ziv of the Technion Institute in Haifa Israel. Here we are using time-lapse confocal imaging of GFP-tagged synaptic proteins expressed in cultured hippocampal neurons to follow the time course of synapse formation and the exchange kinetics of proteins under different conditions. Our on going studies have shown that nascent synapses form in about an hour with the presynaptic active zone forming before the postsynaptic density (PSD) (Brelser et al., 2004). Furthermore, we have found that many of the pre and postsynaptic proteins use vesicular intermediates for their delivery to nascent and mature synapses (Zhai et al., 2001; Shapira et al., 2003). Importantly, we find that proteins within synapses are quite dynamic exchanging on the order of 10s of minutes (Tsuriel et al., 2006). This is in stark contrast to the life of an individual synapse that can exist for many hours to days. Our future studies are designed to relate the dynamics of individual proteins to their proposed functions.
Synaptic Plasticity
Neurodevelopmental Disorders
A new direction for the laboratory has been to link changes in synaptic plasticity to deficits in cognitive ability in individuals with inborn genetic lesions. Current efforts are directed towards understanding the root causes of cognitive impairment in individuals with Down syndrome and whether treatment strategies can be developed to normalize cognitive performance. To accomplish these goals, we study the characteristic of several mouse models of Down syndrome including the Ts65Dn mouse created by Murial Davidson and the Ts1Cje mouse made by Charlie Epstein. Specifically, we have tried to address three questions. First, do the nervous systems of these animals develop normally. For example, do neurons differentiate and form synaptic contacts at normal rates and numbers? A related question is whether the circuit properties of neuronal networks are altered in these mice and at what developmental stage does this begin to appear? Our ongoing studies indicate that neuronal differentiation and synaptogenesis in normal in these mice, yet the circuit properties of neuronal networks are subtly altered such that normal synaptic plasticity mechanisms are impaired (Hansen et al., 2007). Of particular note, we find that the balance of excitation and inhibition in the brains of these mice is shifted such that there is too much inhibition.
These studies have lead to new questions including whether these changes in neuronal excitability and synaptic plasticity lead to alterations in behavior and cognitive performance in these mice and/or whether drugs that reduce inhibition can lead to a normalized cognitive performance. In an elegant study by Fabian Fernandez, a graduate student in the laboratory, we have found that the answer to both of these questions is yes. Specifically, he has found that Ts65Dn mice, similar to humans with Down syndrome, have reduced hippocampal dependent cognitive abilities. Moreover, he has found that by reducing inhibition in the brains of these mice, with non-competitive GABAA receptor antagonists, Ts65Dn mice can perform a variety of memory tasks at a similar level to wild type mice (Fernandez et al., 2007). These results suggest that by subtly reducing inhibition in individual with Down syndrome, with these or related drugs could have some therapeutic benefit for this patient population (Fernandez and Garner, 2007).
Clearly translating finding in mice to humans is a challenging endeavor and can take years to achieve. To make this dream a reality, we have expanded our Down syndrome research program to test the safety and efficacy of a number of different drugs in mice with the vision of moving the best and safest drugs into the clinic.
The work described here was supported by grants from the National Institutes of Health, the Nancy Pritzker Foundation, the Hillblom Foundation, the Down syndrome research and treatment foundation (dsrtf.org), the Deutsche Forschungsgemeinschaft (DFG) and US-Israel Binational Science Foundation (BSF) .
