Investigating the Mitochondrial Bioenergetic Responses to Nanosecond Pulse Stimulation
Abstract/Description/Artist Statement
Background: Nanosecond Pulsed Electric Fields (nsPEF) induce depolarization of the inner mitochondrial membrane (ΔΨm), an increase in the production of mitochondrial reactive oxygen species (mROS), as well as changes in ion and nucleotide gradients. Despite these findings, the regulatory mechanisms governing these responses remain unclear.
Hypothesis: This study proposes that the responses to the nsPEF’s observed are at least partly modulated by the mitochondrial uncoupling proteins (UCP’s) with the possibility of either upstream or parallel responses by energetic stress signaling pathways like the AMP-activated protein kinase (AMPK).
Methods: H9c2, rat cardio myoblasts, and Jurkat E6-1, T-cell lymphoblasts, were exposed to nanopulse stimulation (NPS) in the presence of Adenosine Diphosphate (ADP) and Guanosine Diphosphate (GDP) and the pharmacologic regulators of the UCP genipin and oleic acid. Flow cytometry was used to observe changes in ΔΨm and mROS levels at given time intervals post-nsPEF.
Results: nsPEF exposure produced a dose-dependent decrease in ΔΨm and an increase in mROS. ADP greatly limited these responses, and GDP showed similar but smaller results. genipin and oleic acid showed a great modulation of these changes as well, supporting the involvement of some UCP-mediated Conductance.
Conclusion: These findings support a nucleoside-sensitive, UCP-linked mechanism regulating mitochondrial responses to nsPEFs. Continuing studies will focus on UCP and p-AMPK expressions to determine whether AMPK may participate in parallel, or even upstream, to coordinate these mitochondrial responses. This includes protein quantification and investigation of these effects in knockdown models to demonstrate the mitochondrial reliance on these pathways.
Key Words:
Nanosecond Pulsed Electric Field (nsPEF), Inner Mitochondrial Membrane (IMM), Mitochondrial Uncoupling Proteins, AMP-activated Protein Kinase (AMPK), Inner Mitochondrial Membrane Potential (ΔΨm), Mitochondrial Reactive Oxygen Species (mROS)
Faculty Advisor/Mentor
Stephen J. Beebe
Faculty Advisor/Mentor Email
sbeebe@odu.edu
Faculty Advisor/Mentor Department
Frank Reidy Research Center for Bioelectrics
College/School Affiliation
Other
Student Level Group
Graduate/Professional
Presentation Type
Oral Presentation
Investigating the Mitochondrial Bioenergetic Responses to Nanosecond Pulse Stimulation
Background: Nanosecond Pulsed Electric Fields (nsPEF) induce depolarization of the inner mitochondrial membrane (ΔΨm), an increase in the production of mitochondrial reactive oxygen species (mROS), as well as changes in ion and nucleotide gradients. Despite these findings, the regulatory mechanisms governing these responses remain unclear.
Hypothesis: This study proposes that the responses to the nsPEF’s observed are at least partly modulated by the mitochondrial uncoupling proteins (UCP’s) with the possibility of either upstream or parallel responses by energetic stress signaling pathways like the AMP-activated protein kinase (AMPK).
Methods: H9c2, rat cardio myoblasts, and Jurkat E6-1, T-cell lymphoblasts, were exposed to nanopulse stimulation (NPS) in the presence of Adenosine Diphosphate (ADP) and Guanosine Diphosphate (GDP) and the pharmacologic regulators of the UCP genipin and oleic acid. Flow cytometry was used to observe changes in ΔΨm and mROS levels at given time intervals post-nsPEF.
Results: nsPEF exposure produced a dose-dependent decrease in ΔΨm and an increase in mROS. ADP greatly limited these responses, and GDP showed similar but smaller results. genipin and oleic acid showed a great modulation of these changes as well, supporting the involvement of some UCP-mediated Conductance.
Conclusion: These findings support a nucleoside-sensitive, UCP-linked mechanism regulating mitochondrial responses to nsPEFs. Continuing studies will focus on UCP and p-AMPK expressions to determine whether AMPK may participate in parallel, or even upstream, to coordinate these mitochondrial responses. This includes protein quantification and investigation of these effects in knockdown models to demonstrate the mitochondrial reliance on these pathways.
Key Words:
Nanosecond Pulsed Electric Field (nsPEF), Inner Mitochondrial Membrane (IMM), Mitochondrial Uncoupling Proteins, AMP-activated Protein Kinase (AMPK), Inner Mitochondrial Membrane Potential (ΔΨm), Mitochondrial Reactive Oxygen Species (mROS)