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My research career has
been primarily concerned with elucidating the
mechanisms responsible for regulating blood flow. In
the 1970s and 1980s, my research focused on the role
of adenosine in regulating blood flow, particularly
in skeletal muscle. In the late 1980s, my research
moved away from adenosine when began to examine the
role of nitric oxide (NO) as a regulator of cardiac
and vascular function. In 1986, NO was identified as
an important factor released by vascular endothelium
that causes vasodilation through the cGMP pathway.
Subsequent to that discovery, NO was found to be
produced by most cells within the body, and has been
identified as having a number of important
physiological actions including vasodilation,
inhibition of platelet aggregation, depression of
myocardial contractility, and modulation of neuronal
activity in both the central and peripheral nervous
systems. Nitric oxide, however, is not alone as an
important regulator of vascular function. We now
know that NO has important interactions with other
vascular control mechanisms, including endothelin-1
(ET-1), reactive oxygen species (ROS) and alpha-adrenoceptors. |
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For the past few years,
my research has increasingly concentrated on
interactions between NO, ET-1 and alpha-adrenoceptors.
Initially, I examined these interactions in models
of endotoxemia and septic shock; however, over the
past 3 years my research on vascular control
mechanisms has moved into the field of diabetes. The
following summarizes in more detail some of work
recently completed or currently being conducted in
my laboratory.
Nitric oxide and
endothelin modulation of norepinephrine-induced
coronary vasoconstriction
 This
project, conducted in the summer of 2003, used
isolated, Langendorff-perfused rat hearts to study
how NO and ET-1 modulate norepinephrine (NE)-induced
vasoconstriction in the coronary circulation.
Kristin Prock is presently a second year medical
student and Megan Porter is a third year medical
student and at OUCOM. This work is summarized in the
following abstract that was presented at the
Experimental Biology 2004 meeting held in Washington
D.C.:
Nitric oxide
and endothelin modulation of norepinephrine-induced
coronary vasoconstriction in isolated rat
hearts. Megan E. Porter, Kristin A. Prock,
Richard E. Klabunde. Department of Biomedical
Sciences, Ohio University College of Osteopathic
Medicine, Athens, OH 45701.
Coronary vascular function is regulated, in
part, by endothelial-derived factors such as
nitric oxide (NO), a vasodilator, and endothelin
(ET-1), a vasoconstrictor. The purpose of this
study was to determine how these vasoactive
compounds modify norepinephrine (NE)-induced
vasoconstriction in the beating rat heart.
Hearts from male Sprague Dawley rats were
isolated and perfused by the Langendorff method
with a Krebs Henseleit buffer under constant
flow conditions. L-NAME (NO synthase inhibitor),
BQ-123 (ETA receptor antagonist), propranolol
(β-adrenoceptor blocker), or nothing was added
to the buffer reservoir 40 minutes prior to
infusing NE into the coronaries at
concentrations of 10-9 - 10-5 M. NE alone did
not cause vasoconstriction; however, NE
constricted the coronaries in the presence of
L-NAME. Adding BQ-123 to the L-NAME reversed the
NE-induced vasoconstriction at high NE
concentrations (10-6 and 10-5 M), causing vasodilation. Propranolol added to the L-NAME
and BQ-123 reversed the vasodilation at 10-6 M
NE (p<0.05) and decreased the magnitude of
dilation at 10-5 M. We speculate that the
vasoconstriction observed in the presence of
L-NAME is a result of an absence of NO, which
antagonizes NE vasoconstrictor actions, as well
as an increase in the production of ET-1 caused
by NE. These data also suggest that the vasodilation observed in the presence of BQ-123
is due to NE binding to vascular β2 receptors.
Endothelin
receptors in the microcirculation
This project is
examining the interactions between NO and ET-1 in
arterioles located in the rat cremaster muscle by
using intravital video-microscopy. Kristin Prock and
Sara Klug , both second year OUCOM medical students,
worked on this project in the summer of 2004. This
work followed an earlier study by John Ross, who is
now a first year OUCOM medical student. The
following is an abstract that will be presented at
the Experimental Biology 2005 meeting to be held in
San Diego in April 2005.
Functional
role of endothelin receptors in the rat
cremaster muscle microcirculation. Sara E.
Klug, Kristin A. Prock, Richard E. Klabunde:
Department of Biomedical Sciences, Ohio
University College of Osteopathic Medicine,
Athens, OH 45701
 Endothelin-1
(ET-1) has been implicated in vascular
dysfunction associated with hypertension and
diabetes. ET-1 produces vasoconstriction through
binding to vascular smooth muscle (VSM) ETA
and ETB receptors; ET-1 binding to
endothelial ETB receptors produces
vasodilation through nitric oxide (NO)
formation. Therefore, endothelial ETB
receptor stimulation can modify the effects of
VSM ETA and ETB receptor
stimulation. The purpose of this study was to
determine the role of ETA and ETB
receptors in arteriolar responses to ET-1 in the
rat cremaster muscle microcirculation. The
cremaster muscle of anesthetized rats was
exteriorized and superfused with a Krebs-Henseleit
buffer. Arterioles (~50 µ diameter) were
visualized by intravital videomicroscopy and
diameters measured using a video caliper. BQ-123
(ETA antagonist) produced arteriolar
vasodilation indicating that basal ET-1 release
was influencing vascular tone through ETA
receptors; BQ-788 (ETB antagonist)
caused only slight vasodilation. Both BQ-123 and
BQ-788 effectively blocked the vasoconstriction
produced by ET-1 (10-10 M). The NO
synthase inhibitor, L-NA, did not alter ET-1
vasoconstriction. Therefore, in the rat
cremaster muscle, ET-1 produces arteriolar
vasoconstriction through both ETA and
ETB receptors on the VSM. There was
no evidence for endothelial ETB
receptor modulation of ET-1 vasoconstrictor
responses through NO formation.
Mechanisms of
coronary dysfunction in obesity-induced diabetes
Shawn Bender, a Ph.D.
student, is studying the effects of obesity-induced
diabetes on coronary vascular function in mouse
hearts. Mice are placed on a high-fat,
high-carbohydrate diet for 15 weeks, which produces
significant obesity and hyperglycemia. The mouse
hearts are removed and the coronary vessels perfused
by the Langendorff method. Shawn's dissertation work
is directed toward studying how obesity-induced
diabetes alters the interactions between NO, ET-1
and alpha-adrenoceptors. Shawn has been assisted in
his research by Nathan Lott, a second year OUCOM
medical student, and by Emily Drummond, a research
technician. This work is summarized below in an
abstract that will be presented at the Experimental
Biology 2005 meeting to be held in San Diego in
April 2005.
Diet-induced
obesity alters nitric oxide, endothelin-1 and
alpha-adrenoceptor responses in isolated mouse
hearts. Shawn Bender, Nathan Lott, Richard
Klabunde: Department of Biomedical Sciences,
Ohio University College of Osteopathic Medicine,
Athens, OH 45701.
 Diet-induced
obesity and the metabolic syndrome are major
risk factors for the development of vascular
diseases such as atherosclerosis and
hypertension. The progression of these diseases
is characterized by endothelial dysfunction
leading to altered responses to vasoactive
substances including nitric oxide (NO) and
endothelin-1 (ET-1) as well as to α-adrenoceptor
(α-AR) activation. Microvascular studies as well
as those using isolated heart models have shown
that these conditions result in enhanced
vasoconstrictor responses associated with
reduced NO bioavailability. The purpose of this
study was to characterize the response of the
isolated heart from control and obese mice to
L-NAME (NOS antagonist), ET-1, and phenylephrine
(PE, α1-AR agonist). Hearts were
rapidly excised, cannulated, and perfused under
constant flow conditions with a Krebs-Henseleit
buffer to which all drugs were added at known
concentrations. Coronary perfusion pressure was
measured through a side port in the cannula.
L-NAME produced an approximately two-fold
greater increase in coronary vascular resistance
in control mice than in obese mice. Also, PE and
ET-1-mediated constriction was not present
unless NO production was blocked with L-NAME in
control and obese mice. Following L-NAME
treatment, control mice showed an enhanced
PE-induced constriction versus obese mice.
Therefore, in the isolated mouse heart, PE and
ET-1-mediated constriction is normally masked by
NO-mediated vasodilation. Also, diet-induced
obesity results in attenuated NO bioavailability
and reduced α-AR constriction versus controls.
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