Measurement of ATP-Induced Membrane Potential Changes in IVD cells

Area of Science:

• Biochemistry
• Cell Biology
• Physiology

Background:

• Extracellular adenosine-5'-triphosphate (ATP) is a signaling molecule affecting cell membrane potential and excitability.
• Compressive loading of intervertebral disc (IVD) cells increases ATP production and release, leading to its accumulation in the nucleus pulposus (NP).

Purpose of the Study:

• To develop a noninvasive system for measuring ATP-induced changes in porcine IVD cell membrane potential.
• To investigate the responses of nucleus pulposus (NP) and annulus fibrosus (AF) cells to extracellular ATP in various culture conditions.

Main Methods:

• Utilized a potential-sensitive dye (di-8-ANEPPS) to measure membrane potential changes in response to ATP.
• Examined IVD cell responses in both monolayer and 3D cultures.
• Assessed gene expression of P2X4 purinergic receptors and used a P2 receptor inhibitor (PPADS) to block ATP-induced responses.

Main Results:

• ATP induced dose-dependent changes in membrane potential of porcine IVD cells.
• The magnitude and pattern of these changes varied based on cell type (NP vs. AF), culture condition, and ATP concentration.
• P2X4 purinergic receptor gene expression was detected in both cell types, and PPADS inhibited the ATP-induced responses.

Conclusions:

• Extracellular ATP modulates the biological activities of IVD cells, specifically NP and AF cells.
• These modulations occur through P2 purinergic receptors.
• The cellular response to ATP is complex and influenced by multiple factors including cell type and culture environment.