M.B.A., University of New South Wales (AGSM), Australia, 1991
Ph.D., University of Sydney, Australia, 2000
and pH on the release of neurotransmitters.
Rapid communication between cells is the basis for most of the functions of the nervous system, including sensory transduction, learning, memory, and locomotion. Most cells of the nervous system communicate at synapses, specialized sites at which closely opposed cells release chemicals known as neurotransmitters. Our laboratory investigates the release of neurotransmitters from nerve cells and factors that influence their release, such as mitochondria, pH, and age. We use electrophysiology and high-speed fluorescence imaging to construct a molecular, biophysical, and physiological understanding of neurotransmitter release. We conduct our studies primarily in the fruit fly because of its molecular genetic advantages, but we also conduct studies in mice to examine the effects of mitochondrial dysfunction and age on neurotransmitter release in a vertebrate nervous system.
Project 1: Mitochondria (funded by the National Institute of Health (NIH)).
Mitochondria generate most of the ATP required for the release and recycling of neurotransmitters. They also play an important role in the regulation of Ca2+, an ion which triggers the release of neurotransmitters. Mitochondrial health and tight regulation of neuronal Ca2+ levels are essential as mitochondrial dysfunction and Ca2+ dysregulation are hallmarks of many neurodegenerative diseases.
We have developed cutting edge imaging techniques to investigate the ways in which mitochondria contribute to neurotransmitter release and Ca2+ regulation. We have recently elucidated some of the cellular processes that coordinate mitochondrial ATP production with the energy demands of neurotransmitter release and Ca2+ regulation (Chouhan et al., 2012, J Neurosci, 32, p.1233). We welcome the contribution of graduate students interested in investigating the mechanisms that coordinate mitochondrial ATP production with the energy demands of neurotransmission, and the mechanisms that distribute mitochondria to sites of release according to the energy demands at those sites.
Figure 1. In nerve terminals, mitochondrial energy metabolism increases in response to nerve stimulation; both pH of the mitochondrial matrix (pHm), and the potential of the inner mitochondrial membrane (ΔΨm), increase.
A, Schematic of a motor neuron terminal examined in situ. B, A nerve stimulus train (bar: 80 Hz, 2 s) evoked changes in the concentration of cytosolic and mitochondrial Ca2+ [Ca2+]c) and [Ca2+]m) reported by different Ca2+-indicators in the cytosol (GCaMP3) and matrix (rhod-FF). C, Nerve stimulus trains also evoked changes in pHm, reported by a pH-indicator (mtAlpHi). D, Nerve stimuli also evoked changes in ΔΨm, reported by TMRE. Adapted from Chouhan et al. (2012).
Project 2: pH (funded by the National Science Foundation (NSF)).All biochemical processes are sensitive to the proton concentration (pH) of the environment in which they occur, and dysregulation of neuronal pH is thought to contribute to cell death following hyperexcitable states (epilepsy) and ischemic events (stroke). Recently, our laboratory engineered several genetically-encoded fluorescent pH indicators and used these reporters to determine that the cytosolic pH within nerve terminals changes substantially during nerve activity in situ and in vivo. This observation has profound implications for neurotransmitter release and the mechanisms that underlie the modulation of transmitter release over time (synaptic plasticity) as all these processes rely upon pH-sensitive enzymatic reactions. A more thorough understanding of the effects of pH changes on synaptic transmission will yield insights into the cellular mechanisms of learning and memory as well as the pathology of epilepsy and the ability of the nervous system to recover after a stroke.
We would welcome the contribution of graduate students interested in the following projects: First, we are developing optogenetic tools to control the pH of individual neurons in situ and in vivo. Secondly, we wish to use this technology to determine the influence of pH changes on several neurotransmitter release processes (e.g. Ca2+ buffering, synaptic vesicle endocytosis and loading) and the extent to which they influence synaptic plasticity.
Figure 2. Motor activity causes substantial acidification in a motor nerve terminal. Traces representing simultaneous changes in cytosolic Ca2+ levels and pH ([Ca2+]cyto and pHcyto respectively), reported by rhod-dextran and superecliptic pHluorin, in response to nerve stimulus trains (black bars, 40Hz, 2s). Rossano, Chouhan and Macleod - unpublished.
Project 3: Age (funding sought from the National Institute of Aging (NIA) & Muscular Dystrophy Association (MDA)).
The performance of the nervous system and the behaviors for which it is responsible decline with age. In collaboration with the laboratory of Dr. Benjamin Eaton we found that fruit flies show motor deficits with age (Rawson et al., 2012, Aging Cell, 11, p.418) along with changes in neurotransmitter release from motor neurons. In collaboration with Dr. Holly van Remmen, we have found that mice show mitochondrial Ca2+ handling deficits in motor neurons with age. We would welcome the contribution of graduate students interested in investigating the impact of age-related changes in mitochondrial Ca2+ handling and the progression of certain diseases that affect the release of neurotransmitters from motor neurons, such as amyotrophic lateral sclerosis (ALS).