For proper neuron function, the neuron must have the correct number of input dendrites, which look like branches on a tree. Very little is currently known about how the pattern of these branches is determined or how these branches change when a person learns. To make new dendrite branches, a cytoskeleton must be produced, much like the skeleton in fingers. Dr. Firestein discovered a protein called cypin that plays a critical role in making this cytoskeleton, and hence dendrites. It is hypothesized that without cypin, neurons will not form properly, and the brain will not function correctly. Using nerve cells in a dish, undergraduate and graduate students will perform experiments to understand the mechanism by which cypin acts to yield functioning neurons. By investigating how cypin gives nerve cells their shape and how these nerve cells integrate into simple circuits, this research will aid in our understanding of principles by which circuits may be modified during events, such as learning. Dr. Firestein and colleagues will include a diverse group of undergraduate and graduate students and will establish an exchange program with University of Puerto Rico. This proposal also encompasses activities to excite a younger generation of students (K-12) about neuroscience by producing a video series "Teach Me Neuroscience" and training K-12 teachers at the bench. It is Dr. Firestein's hope to establish a program to bring neuroscience to the community in Puerto Rico via seminars, workshops, and exchange programs.
The specific goal of the current work is to evaluate how cypin and its binding partner PSD-95 affect neural circuit dynamics. Experiments aim to determine the mechanism by which cypin decreases synaptic PSD-95 using viral-mediated gene expression in cell culture to alter cypin levels. The role of the proteasome in cypin-mediated changes in PSD-95 will be assessed. It will also be determined whether cypin and PSD-95 levels affect function and activity of neural circuits in vitro. Dr. Firestein has a comprehensive set of tools available to manipulate cypin functionality at the molecular level and will assess effects of altered cypin levels on dendrite number, spine number, and size. Experiments will make use of electrophysiology techniques to determine whether cypin affects neuronal signaling. The proposed work uses interdisciplinary approaches - molecular/cellular, biochemical, and electrophysiological - to understand how morphological changes to neurons result in changes in synaptic function, making a large advance from previous cell culture work and defining a mechanism by which cypin acts to regulate dendritogenesis and determine effects on neural circuitry.
