Our lab studies the mechanisms of calcium deregulation, mitochondrial dysfunction, and neuronal loss in aging and age-related neurodegenerations.

We use transgenic and knock-in mouse models of Huntington’s disease and cyclophilin D-knockout mice    (Ppif ^-/-) to study the mechanisms of Huntington’s disease (HD) and the role of calcium deregulation and mitochondrial injury in HD progression. 

We utilize highly purified isolated brain (synaptic and non-synaptic) mitochondria to analyze the effect of aging on mitochondrial functions. To study isolated mitochondria, we use a computerized setup for simultaneous measurements of respiration, membrane potential, swelling, and calcium uptake by mitochondria. Using isolated mitochondria, we study the mechanisms of mitochondrial injury in aging, particularly, the role of the mitochondrial permeability transition pore. We also investigate the effect of aging on reactive oxygen species (ROS) production in mitochondria.

In addition to isolated mitochondria, we use cultured neurons derived from newborn and adult mice and rats. We use cellular respirometry (Seahorse technology), live-cell fluorescence wide-field and a laser spinning-disk confocal microscopy to investigate mitochondrial dysfunction, calcium deregulation, and ROS production in cultured neurons. We use electron microscopy to analyze mitochondrial morphological changes. We employ western blotting and immunocytochemistry to evaluate expression of different proteins and investigate the release of mitochondrial proteins involved in programmed cell death, apoptosis.

The overall goal of our lab is to determine the mechanisms of calcium deregulation, mitochondrial dysfunction, and neuronal death in aging and age-related neurodegenerations such as Huntington’s disease.

Figure 1.  Simultaneous measurements of respiration and membrane potential Δψ_m with isolated rat brain mitochondria.

Figure 2.  Recordings of mitochondrial membrane potential in individual isolated brain mitochondria loaded with a fluorescent dye Rhodamine 123.  A, fluorescence image of individual mitochondria attached to glass coverslip.  B, fluorescence traces showing normalized (F/F₀) Rhodamine 123 signal indicative of mitochondrial polarization.

Figure 3. The 3D reconstruction of mitochondrial network in live cultured hippocampal neurons.

Figure 4.  The 3D reconstructions of individual mitochondrion in live hippocampal neuron exposed to glutamate.

Figure 5.  Oxygen consumption by cultured hippocampal neurons measured with Seahorse XF24 analyzer.

Figure 6.  Glutamate-induced calcium deregulation in neurons from 8-month-old FVB/NJ mice. 
a. immunostaining with anti-MAP2 antibody (green) to visualize neurons and staining with DAPI to show nuclei (blue). The large nuclei outside of neurons belong to glial cells. b-e, selected pseudocolor images of an adult mouse (8 month old) neuron (10 days in vitro) loaded with the Ca²⁺-sensitive dye Fura-2FF and treated with 10mM glutamate plus 10mM glycine. The images were taken at indicated time points. f, fluorescence traces from three individual adult neurons in the typical experiment are shown. Arrowheads on the red trace indicate when images shown in b-e were taken.