“Effect
of Mitochondria on Calcium Homeostasis in Neuronal Cells”
Student Project: Nasibo Kadir, leogrrl21@hotmail.com
Statement of Project: This
is my summary of recent research in the field of mitochondrial-associated
aging, based on the research article, “Changes in Mitochondrial Status
Associated with Altered Ca²+ Homeostasis in Aged Cerebellar Granule
Neurons in Brain Slices,” Jie Xiong, Alex Verkhratsky, and Emil C.
Toescu, Journal of Neuroscience (2002), 22(24): 10761-10771.
This is my project for BIOL
475, Neurobiology, Dr. Grant Mastick, Biology Department,
INTRODUCTION:
● For
centuries, human beings have struggled with aging and have looked for and
developed methods to slow it down and/or reverse it. This includes creams,
medicines, hi-tech procedures, and cosmetic surgery, among others.
● In
the last few decades, scientists have studied the phenomenon of aging at the
cellular level, and many have now concluded that the physical and mental
changes associated with aging are connected to mitochondrial function within
each respective cell in the body.
● Mitochondria are the
“powerhouse of the cell.” This is the organelle that provides
energy for all cellular functions by generating ATP (adenosine triphosphate),
and there are thousands of them in each cell in order to meet the cell’s
energy demands. Thus, mitochondrial defects tend to cause major problems within
the cell itself. Mitochondrial defects are caused when damage occurring in
mitochondrial DNA (mtDNA) causes mutations to add up within the mitochondria.
This in turn leads to the damage that scientists believe actually causes aging
in cells, tissues, and eventually in the whole organism. These mitochondrial
defects can be caused by:
▪ build-up of reactive oxygen species (ROS) that are
byproducts of cellular respiration,
▪ disturbances in calcium homeostasis; excessive calcium
within mitochondria causes damage, and thus Ca²+ has to be carefully
regulated,
▪ etc.
Hypothesis of this
particular study: There is a relationship
between mitochondrial function and Ca²+ homeostasis in old versus young
neuronal cells.
cerebellum
Figure 1. An image of the mammalian brain (human).
EXPERIMENTAL SYSTEM:
Experiment 1:
In order to test their
hypothesis, the researchers took cerebellar slices from both young (6-8 mos.)
and old (20-24 mos.) mice in order to compare Ca²+ homeostasis between
them. The slices were placed in solution and then mounted on an upright
microscope and then stimulated. Images were then taken of the regions of
interest, which were the soma of the neurons. Using the standard Grynkiewicz
formula, Ca²+ concentration levels were determined 1-1.5 hours and 3 hours
after slicing.
Experiment 2:
The researchers also wanted to observe any changes in Ca²+ concentration between both the young and old neurons after stimulation, particularly how long it would take the Ca²+ concentration within the cells to return to their resting levels. To achieve this, the cell membrane of the respective cells was stimulated via KCl-evoked depolarization using 50mM KCl.
RESULTS:
Images taken at 1-1.5 hours
after slicing showed no significant change in Ca²+ concentration between
the old and new cerebellar slices. The Ca²+ concentration between the two
did differ however 3 hours after slicing. The researchers found that there was
a higher Ca²+ concentration in the old cells than in the new cells after 3
hours, which led them to conclude that the old cells had a more difficult time
returning to the resting Ca²+ concentration than the young cells.
Figure 2. Shows Ca²+ concentration in both young and old cells from beginning
of
experiment until 6 hours after slicing. There is a negligible
difference in Ca²+ concentration
between the old and young slices up until 1.5 hours after
slicing. After 3 hours, the young
cells show a lower Ca²+ concentration, indicating
that there was a quicker return to the
resting Ca²+ concentration (Xiong et al. 2002).
In the second experiment
concerning depolarization with KCl, the researchers found that in the young
brain slices, almost all of the neurons were able to achieve full recovery of
the resting Ca²+ concentration. In the old slices, some of the neurons
were able to do the same, but the remaining either responded to depolarization
but were unable to fully return to the resting state concentration, or they did
not respond to the depolarization at all and had a Ca²+ concentration that increased instead (see figures below).
Figure 3. Depolarization-induced Ca²+ responses in
cerebellar granule neurons of young
mice. There is a definite depolarization and a subsequent
repolarization as the membrane
returns to normal resting state (Xiong et al. 2002).
Figure 4. Depoalrization-induced Ca²+ response in cerebellar granule neurons of old mice.
There is a depolarization,
but there is a failure to repolarize, and the membrane cannot return
to its resting state (Xiong et al. 2002).
Figure 5. Depolarization-induced Ca²+ response in cerebellar granule neurons of old mice.
There is no depolarization
in response to the KCl stimulation, and the Ca²+ concentration
levels actually increased in neurons with this type of
response (Xiong et al. 2002).
CONCLUSIONS:
Based on the results of the
above experiments, the researchers concluded that the delay in the recovery of
the resting Ca²+ concentration found in some of the neuronal cells
was linked to some sort of mitochondrial defect or impairment since
mitochondria are responsible for the regulation of Ca²+ levels. They also concluded that this mitochondrial impairment was
linked to aging, because the old neurons were much more affected than the young
ones.
SIGNIFICANCE:
The results yielded by this
experiment are significant in that they provide strong evidence showing a link
between mitochondrial function and Ca²+ homeostasis, and
between mitochondrial defect and cellular aging.
FUTURE DIRECTION
Using the information gained
in this experiment and from many others, Science may one day be able to more
fully understand the role of mitochondria in aging, particularly its effect on
neurons. From here, it is hopeful and probable that researchers will be able to
develop treatments, and perhaps cures, for some of the neurodegenerative
diseases out there that develop during old age and that are thought to be
caused by mitochondrial defects (i.e., Alzheimer’s, Parkinson’s
Disease, etc.). For now, more research is required in this field.
REFERENCES:
Primary research
article: “Changes in Mitochondrial
Status Associated with Altered Ca²+ Homeostasis in Aged Cerebellar Granule
Neurons in Brain Slices,”
Xiong, J., A. Verkhratsky, and E.C. Toescu, Journal of Neuroscience
(2002), 22(24): 10761-10771. link to article.
Review article: “Mitochondrial Oxidative Stress Plays a Key Role
in Aging and Apoptosis,” Sastre, J., F.V., Pallardó, and J.
Viña, IUBMB Life (2000), 49:
427-435. link to article.
Related articles:
“Calcium Homeostasis and Reactive Oxygen
Species Production in Cells Transformed by Mitochondria from Individuals with
Sporadic Alzheimer’s Disease,”
Sheehan, J.P., R.H. Swerdlow, S.W. Miller, R.E. Davis, J.K. Parks, W.D.
Parker, and J.B. Tuttle, Journal of Neuroscience (1997), 17(12):
4612-4622. link to article.
“Aging: Mice and
mitochondria,” Martin, G.M. and L.A. Loeb, Nature (2004), 429(6960):
357-358. link to article.
This page was constructed by:
Nasibo Kadir, leogrrl21@hotmail.com,
November, 16 2004.