Motor neuron (MN) diseases such as spinal muscular atrophy and amyotrophic lateral sclerosis lead to progressive degeneration of MNs, presenting first with muscle weakness, followed by locomotor defects and frequently death due to respiratory failure. While progress has been made in identifying genes associated with MN degeneration, the molecular and cellular processes underlying disease onset and progression remain unclear, and no effective therapies are available. Methods to direct the development of normal and diseased motor neurons from human embryonic and induced pluripotent stem cells have recently been developed, raising hope that these cells could offer a means for investigating the root causes of MN disease and devising screens for neuroprotective agents. Most stem cell-based disease modeling efforts have thus far focused on the issue of MN survival at the end stages of disease progression. However, studies in animal models and human patients indicate that MN function declines well before MN death is prevalent. We have developed a simple, yet physiologically relevant platform for measuring the activity of normal and diseased human MNs and muscle cells in a manner that has not previously been possible. Here we propose to explore how MN function declines; eventually we hope to test new therapeutics. These studies provide a crucial bridge between studies of motor circuit function in animal models and the molecular and cellular tools available to study cells in culture.
Neurological diseases are among the most debilitating medical conditions that affect millions of Californians each year, and many more worldwide. Few effective treatments for these diseases currently exist, in part because we know very little about the mechanisms underlying these conditions. Through the use of human embryonic stem cell and induced pluripotent stem cell technologies, it is now possible to create neurons from patients suffering from a variety of neurological disorders that can serve as the basis for cell culture-based models to study disease pathologies. Our proposed research specifically seeks to develop an innovative system for investigating the early stages of neuromuscular disease onset and progression in an experimentally accessible cell culture setting. The generation of this model will constitute an important step towards understanding the root cause of neurological dysfunction and developing a platform for the discovery of drugs that can alter disease outcomes and improve the productivity and quality of life for many Californians. Moreover, progress in this field will help solidify the leadership role of California in bringing stem cell research to the clinic, and stimulate the future growth of the biotechnology and pharmaceutical industries within the state.
Motor neuron diseases such as Spinal Muscular Atrophy (SMA) or Amyotrophic Lateral Sclerosis (ALS) are associated with progressive degeneration of motor neuron function. To date, there are no established in vitro assays for studying the pathophysiology of motor neuron diseases. This Fundamental Mechanisms proposal aims to develop an in vitro stimulation and recording assay for measuring human neuromuscular synaptic function and dysfunction (Aim 1). This assay will then be used to determine the mechanistic basis for neuromuscular synaptic dysfunction using SMA patient-derived cells (Aim 2).
Significance and Innovation
- While the genes responsible for the onset of SMA have been identified, the actual pathophysiological mechanism of motor neuron death in that disease is still unknown. The proposed research is significant in that it offers the potential to uncover that mechanism.
- The co-culture system to be developed is innovative and uses cutting edge methods, such as optogenetics and imaging calcium with genetically encoded calcium indicators.
- The successful development of the co-culture technology platform will enable studies of the mechanism of action of drugs currently prescribed for specific motor neuron diseases, and in addition will enable screening for new drugs.
- Successful use of this technology for understanding the mechanism of SMA will open the door for its use in studying other motor neuron diseases.
Feasibility and Experimental Design
- The preliminary data, primarily from the murine system, are very compelling and support the idea that there are changes in synaptic function in SMA.
- The application is weakened by the proposed use of only one human SMA disease line and one related but not isogenic control human cell line. Due to this limitation the proposed research might lead to the study of an epiphenomenon rather than an actual disease phenotype.
- The budget is well described and appears appropriate for the completion of the proposed work.
Principal Investigator (PI) and Research Team
- PI and the co-PI are both very strong scientists with excellent publication records. Their complimentary areas of expertise are well suited to conduct the proposed research.
- The PI has committed sufficient effort to the project.
Responsiveness to the RFA
- The proposed project is responsive to the RFA in that it uses human stem cell-derived cells to study mechanisms of motor neuron degeneration in SMA.