Physiology Research Program
General statement on research interests:
The theme of our laboratory is understanding the regulation of smooth muscle contraction and relaxation. In particular, we focus on elucidating mechanisms responsible for unique contractile responses of smooth muscle in several vasculatures. Because regulation of blood flow into a variety of organs is essential for their normal function the major projects of the lab, as outlined below, include efforts to understand the physiologic control of vascular tone, molecular determinates of vascular and non-vascular smooth muscle function, as well as the effects of cardiovascular risk factors on vascular function.
The fundamental hypothesis based on the central theme outlined above is that detriments in vascular function are explained by variations in the signaling elements of smooth muscle comprising the target organs. Vascular smooth muscle is distinct from other smooth muscle types and thus, may express a unique complement of receptors and signaling pathways. To address this, we are examining some of the known pharmacologic behaviors of the smooth muscle in the light of new techniques in molecular biology and animal models of vascular dysfunction. Specifically, we are using the variations in vasoconstrictor/vasodilator sensitivity to determine if there are correlative changes in receptor expression or the associated signaling pathways. We have 3 major projects on going in the lab and are a briefly described below.
Airborne particulate matter and the negative impact on vascular reactivity and cardiovascular health:
Recent observations indicate that pulmonary inflammation induced by airborne particulate matter leads to activation of circulating neutrophils and the coronary endothelium, which enhances myocardial inflammation following ischemia reperfusion. Our data indicates that this effect is associated with endothelial modulation of aortic vascular smooth muscle reactivity in a manner consistent with a defect in adenosine control of endothelial-dependent relaxation. The specific hypothesis of this project is that neutrophilic inflammation induced by acute exposure to PM activates the endothelium, leading to an imbalance in signaling through adenosine A1, A2, and A3 receptors. Subsequent inhibition of A2 receptors by A1 and A3 receptor signaling, reduces endothelial-dependent relaxation of vascular smooth muscle, predisposing the myocardium to infarction; and altered adenosine signaling increases the severity of myocardial inflammation subsequent to reperfusion. We use a mouse model for studies of endothelium-smooth muscle interactions and to define the effects of PM exposure on cellular, protein, and molecular targets in the myocardium during ischemia-reperfusion injury, as well as affects on coronary and pulmonary vasculature, myocardial function, and effects of specific adenosine receptor antagonists. This work will provide definitive mechanistic evidence of the link between airborne particulate and cardiovascular disease.
Impact of RhoA/Rho-kinase signaling on penile smooth muscle function and erection:
Our primary goal in understanding the physiologic regulation of penile circulation is examining the role of elements of the Ca2+-sensitization process on maintaining smooth muscle contraction. Traditionally erection, more specifically erectile dysfunction, has focused on the inability to relax the smooth muscle of penile vasculature by means of nitric oxide production. However, the population of men effectively treated by agents which prolong the effect of NO (e.g. Viagra, Lavitra or Cilais) has not resolved all the dysfunction. An emerging concept is the role signaling pathways like the RhoA/Rho-kinase have on augmenting the contractile sensitivity of the penile smooth muscle. Currently we are exploring interactions between various vasoconstrictors (phenylephrine and endothelin-1) with downstream signaling molecules (RhoA/ROCK/RND and PKC) that mediate a sensitizing effect on smooth muscle contractile tone. These studies include analysis of penile vascular hemodynamics, in situ measurements of blood flow/pressure in response to neurally and pharmacologically evoked erection and in vitro studies of cavernosal tissue reactivity.
Effects of Metabolic Syndrome X on urogenital smooth muscle function:
Several cardiovascular diseases endanger the health of the American people; "Syndrome X" is the most rapidly increasing in incidence. The most widely accepted definition of "Syndrome X" is the combned development of several cardiovascular risk factors including obesity, hypertension, and insulin-resistant (type II) diabetes. A useful animal model with a "Syndrome X"-like phenotype is the Fatty Zucker Rat. Studies currently ongoing in our lab are using this model to determine the nature of urogenital organ disturbances associated with presence of multiple cardiovascular risk factors. Specific focal points in these studies include functional remodeling, changes in the production of nitric oxide and free radicals, and alterations in reactivity to vasoconstrictors in the bladder and corpus cavernosum.
- Animal Care and handling, anesthesia, and maintenance of rat, mouse, and rabbit preparations.
- Surgical preparations for measurements of mean arterial pressure, venous pressure, intra-cavernosal/bladder pressure, aortic and cavernosal blood flow and nerves stimulation and acute/chronic administration of vaso-active compounds in select organ beds and systemically.
- Surgical castration, diabetes, and chronic hormone replacement rodent models.
- Use genetic models of hypertension and obesity/diabetes for bladder and cavernosal function studies.
- Isometric/isotonic tension studies: Force development and shortening in smooth/skeletal muscle and cardiac tissues.
- Dynamic stiffness measurements of isolated smooth muscle tissue using sinusoidal perturbation.
- Fluorescence indicators measurements of Ca2+ using Aequorin and FURA-2 and intracellular pH using BCECF in isolated muscle cells, smooth muscle tissues and vessels, and cardiac strips.
- Energetic measurements: oxygen consumption by Clark-style electrodes and ATPase by NADPH fluorescence.
- Cell Culture: maintenance of smooth muscle cells in culture, isolation of primary cells by explantation and enzymatic dispersion.
- Pharmacological characterization of signal transduction pathways.
- Measurement of protein phosphorylation by 1D and 2D gel electrophoresis.
- Protein detection by western blot analysis fluorescence microscopy of labeled proteins.
- Micro Array and PCR approaches for protein and genomic expression of signaling targets.