The Halpain laboratory works toward understanding molecular mechanisms that underlie neuronal network formation. Our work concerns how the complex morphology of neurons develops, what contributes to the delicate balance between stability and plasticity of form, and what factors underlie destabilization of neuronal structure in the context of disease or injury. Our primary focus is on cytoskeletal proteins, specifically microtubules and actin filaments. We apply a multidisciplinary approach encompassing biochemical and molecular biological techniques, electrophysiology, high resolution light microscopy, time-lapse imaging of living cells, and quantitative image analysis. We use carefully chosen model systems, including, where appropriate, primary neuronal cultures, brain slices, cells lines, and cell-free systems. Our studies are fundamentally relevant to many forms of neurological and neuropsychiatric diseases, including neurodegeneration, stroke, mood disorders, mental retardation and autism.
Early Cytoskeletal Events in Neuritogenesis
An essential early step in the formation of neural networks is the initiation, outgrowth, and polarization of neurons. We have focused on the role of microtubule-associated proteins (‘MAPs’) in these processes. The major conceptual advances derived from our work are that MAP2 acts to coordinate the interaction of microtubules with F-actin during neurite initiation, and that cytoplasmic dynein contributes part of the force needed to push out the plasma membrane during neurite initiation. During the past year we have broadened the scope of our studies on MAPs by incorporating both genomic and proteomic screens, which are leading to the identification of new protein complexes involved in neurite outgrowth and branching.
Regulation of Dendritic Spines
Spines are tiny, actin-rich protrusions along the dendrite that concentrate glutamate receptors and other proteins mediating excitatory synaptic transmission. We investigate mechanisms that regulate spine numbers, shape, and stability. We demonstrated that actin dynamics are critical in the establishment of hippcampal long-term potentiation (LTP), a form of synaptic enhancement that is thought to underlie memory. We also showed that actin filament destabilization is a key mechanism underlying glutamate-induced spine loss, which can occur under certain pathological conditions, such as seizures or CNS trauma. Our work has identified specific molecular regulators of spine actin filaments, the major determinant of spine shape, including the protein phophatase calcineurin and the phospholipid-binding protein MARCKS. Through its signal-dependent membrane association, MARCKS is a key regulator of spine numbers and shape. Currently we are investigating the molecular mechanisms for how MARCKS controls actin assembly and membrane trafficking to regulate spine.
Alzheimer’s Disease and Synaptic Morphology
Soluble oligomeric amyoid beta peptide (A-beta) is a key factor in Alzheimer’s disease pathology. We recently showed that low concentrations of A-beta have profound and rapid effects on synaptic structure and function. Time-lapse imaging of connected pairs of presynaptic and postsynaptic compartments enable us to monitor concurrent changes at both sides of the synapse. Future studies will utilize these and other tools of cell biology to probe mechanisms of synapse dysfunction in Alzheimer’s disease.
Calabrese, B., Shaked, G.M., Tabarean IV, Braga, J., Koo, E.H., and Halpain, S. (2007) Rapid, concurrent alterations in pre- and postsynaptic structure induced by naturally-secreted amyloid-beta protein. Molec. Cell Neuroscience 35: 183-193.
Dehmelt, L., Nalbant, P., Steffan, W., and Halpain, S. (2006) A microtubule-based, dynein-dependent force induces local cell protrusions: implications for neurite initiation. Brain Cell Biology 35: 43-60.
Calabrese, B., Wilson, M., and Halpain, S. (2006) Development and regulation of dendritic spine synapses. Physiology 21: 38-47.
Calabrese, B. and Halpain, S. (2005) Essential role for the PKC target MARCKS in maintaining dendritic spine morphology. Neuron 48: 77-90.
Dehmelt, L and Halpain, S. (2004) Actin and microtubules in neurite initiation: Are MAPs the missing link? J. Neurobiol. 58: 18-33.
Roger, B.*, Al-Bassam, J*., Dehmelt, L., Milligan, R., and Halpain, S. (2004) MAP2c, but not tau, binds and bundles F-actin via its microtubule binding domain. Current Biology 14: 363-371. (*these authors contributed equally)
Dehmelt, L., Smart, F., Ozer, R., and Halpain, S. (2003) The role of MAP2c in the reorganization of microtubules and lamellipodia during neurite initiation, J. Neuroscience, 23: 9479-9490.
Al-Bassam, J., Ozer, R., Safer, D., Halpain, S., and Milligan, R. (2002) MAP2 and tau bind along the outer ridges of microtubule protofilaments. J. Cell Biol. 157: 1187-1196.
Krucker, T., Siggins, G.S., and Halpain, S. (2000) Dynamic actin filaments are required for stable LTP in area CA1 of the hippocampus. Proc. Natl. Acad. Sci. (USA) 97: 6856-6861.
Halpain, S., Hipolito, A., and Saffer, L. (1998) Regulation of F-actin stability in dendritic spines by glutamate and calcineurin. J. Neurosci. 18: 9835-9844.
Barros, C.S., Calabrese, B., Chamero, P., Roberts, A.J., Korzus, E., Lloyd, K., Stowers, L., Mayford, M., Halpain, S., and Müller, U. (2009) The neuregulin-1 receptors ErbB2 and ErbB4 cooperate to promote maturation of dendritic spines in central nervous system. Proc. Natl. Acad. Sci. USA 106 (11): 4507-4512.