The long-term goal of our research is to understand how neural circuits are established during development and how they are refined by sensory experience and altered in disease. We are specifically interested in circuits of the cerebral cortex, a part of the vertebrate brain with a central role in many higher order brain functions, including learning and memory, perception and cognition. Disorders in development and plasticity of cortical circuits are thought to contribute to the etiology of many neurological diseases, including schizophrenia, autism spectrum disorder, and Alzheimer’s disease.
In the mammalian cerebral cortex, most excitatory synaptic connections occur on microscopic protrusions that extend from the neuronal dendritic membrane called dendritic spines. Typically, a spine consists of a bulbous head (volume ~ 0.1 -1 µm 3 ) at the end of a thin neck (diameter ~ 0.1 µm). Spines are highly dynamic during development; they form and retract, elongate and shorten in length, and change in volume and shape. These anatomical changes at spines underlie functional changes in cortical circuits. In the Zito laboratory, we use advanced imaging techniques combined with molecular genetics and electrophysiology to study the morphological and functional development of spine synapses when circuits are forming in the cerebral cortex and when they are modified during learning and in disease.
Mechanisms of dendritic spine outgrowth and functional maturation
The growth of new dendritic spines and the formation of new spine synapses are thought to contribute to the circuit rearrangements that occur during learning. Our lab discovered that local regulation of proteasomal degradation in dendrites as a key regulatory mechanism linking synaptic activity with new spine outgrowth. We also identified the RhoGEF, Ephexin5, as a target of the proteasome in regulating new spine outgrowth. Our data support a model in which Ephexin5 (E5) serves a dual role in spinogenesis, operating both as a check on exuberant spinogenesis and as a necessary factor in promoting activity-dependent spine outgrowth. We are currently testing this model using two-photon imaging of FRET-based biosensors in living neurons. Notably, E5 levels are elevated in the brains of patients with Alzheimer’s disease, which is also associated with dramatic loss of dendritic spines. Results from these experiments will further our knowledge of the molecular mechanisms of learning and have the potential to identify novel therapeutic targets for Alzheimer’s disease.
Mechanisms of dendritic spine stabilization
The activity-dependent long-term growth of existing dendritic spines and stabilization of nascent dendritic spines have been widely accepted as key cellular mechanisms supporting learning. Synaptic strengthening of spiny synapses is highly correlated with size increases in dendritic spines. Our lab found that strong glutamatergic stimulation in a pattern that elicits long-term synaptic strengthening is also capable of increasing the stability of newly formed dendritic spines. We further demonstrated that nascent spine stabilization is inhibited by blocking the NMDA-type glutamate receptor (NMDAR) and by select mutations in the NMDAR that interrupt its interaction with the Ca 2+ -calmodulin dependent kinase, CaMKII. Notably, in mature spines, the CaMKII-NMDAR complex is thought to facilitate both CaMKII catalytic and scaffolding activities, either of which might be important for stabilizing new spine structure. We are currently investigating the molecular mechanisms through which CaMKII promotes nascent spine stabilization. In addition, learning is thought to involve the long-term growth of existing spines as synapses strengthen during the circuit changes associated with learning. Notably, learning in humans is improved when breaks are incorporated into learning sessions. Several labs have found that this need for breaks could be due to a temporary saturation of the circuit plasticity necessary for learning. We are currently using two-photon imaging of FRET biosensors, electrophysiology, and calcium imaging to investigate the mechanisms of this temporary saturation of synaptic plasticity at the single synapse level.
Mechanisms of dendritic spine elimination and spine synapse disassembly
Several labs have identified that spine synapse elimination plays a key role in nervous system plasticity, both when circuits are refined during development and when they are modified during learning, yet the molecular and cellular mechanisms driving spine elimination remain ill defined. Early studies identified that dendritic spine shrinkage and loss is associated with long-term depression (LTD) of synaptic strength. Our lab has shown that this destabilization of dendritic spines during LTD can be driven either by input-specific mechanisms or by local heterosynaptic competition between neighboring synapses on dendritic segments. Intriguingly, we have shown that ion flux-independent NMDAR signaling driven by glutamate binding in the absence of co-agonist is sufficient to drive the dendritic spine shrinkage associated with LTD. Furthermore, we have identified p38 MAP kinase, the interaction between NOS1AP and nNOS, nNOS enzymatic activity, activation of the kinase MK2 and cofilin, and signaling through CaMKII as key players in this signaling pathway. Notably, recent studies from our lab have suggested that ion flux-independent NMDAR signaling gates the bidirectional spine structural changes vital for brain plasticity, most likely through destabilizing the actin cytoskeleton in preparation for plasticity. We are currently working toward delineating the molecular mechanisms that link NMDAR conformational changes to spine structural changes and also how these signaling mechanisms combine to drive local competition on dendritic segments.
Error Driven Learning
Error-driven learning is a form of learning in which rapid contrasts between expectations and outcomes are utilized to correct for errors in our expectations. These rapid contrasts allow us to deal with contingencies in our everyday life and learn from situations out of the ordinary. We are currently engaged in a fruitful collaboration investigating the mechanisms of synaptic plasticity in error driven learning with Dr. Randy O'Reilly's laboratory.
With the goal of developing novel glutamate sensors to facilitate our research on activity-dependent spine growth, stabilization and elimination, we teamed up with the labs of Dr. Lin Tian at UC Davis and Dr. Na Ji at Berkeley to develop novel glutamate sensors for large-scale monitoring of the activity of individual synapses in the living behaving animal. In addition, we have teamed up with the lab of Dr. Jon Sack at UC Davis to develop an imaging method utilizing the EVAP (Endogenous Voltage-sensor Activity Probe) tool to for real time measurements of the conformational changes of endogenous ion channel proteins in living tissue.