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Estimation of the membrane potential of cultured macrophages from the fast potential transient upon microelectrode
Abstract:
Analysis of membrane potential recordings upon microelectrode impalement of four types of macrophages (cell lines P388D1 and PU5-1.8, cultured mouse peritoneal macrophages, and cultured human monocytes) reveals that these cells have membrane potentials at least two times more negative than sustained potential values (E(s)) frequently reported. Upon microelectrode entry into the cell (P388D1), the recorded potential drops to a peak value (E(p)) (mean -37 mV for 50 cells, range -15 to -70 mV) within 2 ms, after which it decays to a depolarized potential (E(n)) (mean -12 mV) in about 20 ms. Thereafter, the membrane develops one or a series of slow hyperpolarizations before a final sustained membrane potential (E(s)) (mean -14 mV, range -5 to -40) is established. The mean value of the peak of the first hyperpolarization (E(h)) is -30 mV (range -10 to -55 mV). The initial fast peak transient, measured upon microelectrode entry, was first described and analyzed by Lassen et al. (Lassen, U.V., A.M. T. Nielson, L. Pape, and L. O. Simonsen, 1971, J. Membr. Biol. 6:269-288 for other change in the membrane potential from its real value before impalement to a sustained depolarized value. This was shown to be true for macrophages by two-electrode impalements of single cells. Values of E(p), E(n), E(h), E(s), and membrane resistance (R(m)) measured for the other macrophages were similar to those of P388D1. From these results we conclude that E(p) is a better estimate of the true membrane potential of macrophages than E(s), and that the slow hyperpolarizations upon impalement should be regarded as transient repolarizations back to the original membrane potentials. Thus, analysis of the initial fast impalement transient can be a valuable aid in the estimation of the membrane potential of various sorts of small isolated cells by microelectrodes.
Insights
Macrophage membrane potentials are more negative than previously reported. Analyzing the initial impalement transient, not the sustained potential, provides a more accurate estimate for these cells.
Area of Science:
- Cellular electrophysiology
- Membrane biophysics
- Immunology
Background:
- Accurate measurement of macrophage membrane potential is crucial for understanding cellular function.
- Previous studies often reported sustained potential values (E(s)) that may not reflect the true resting potential.
Purpose of the Study:
- To re-evaluate the membrane potential of various macrophage types using microelectrode impalement.
- To determine the most accurate method for estimating macrophage membrane potential.
Main Methods:
- Microelectrode impalement was used to record membrane potentials in four types of macrophages: P388D1 and PU5-1.8 cell lines, cultured mouse peritoneal macrophages, and cultured human monocytes.
- Analysis focused on the initial peak potential (E(p)) and subsequent potential changes, including sustained potential (E(s)) and hyperpolarizations (E(h)).
- Two-electrode impalements were employed to validate findings in single cells.
Main Results:
- Macrophage membrane potentials were found to be at least two times more negative than previously reported sustained values (E(s)).
- Upon impalement, a rapid peak potential (E(p)) was observed, followed by decay to a depolarized potential (E(n)) and slow hyperpolarizations (E(h)) before reaching E(s).
- The peak potential (E(p)) and transient repolarizations (E(h)) were consistent across all tested macrophage types.
Conclusions:
- The initial peak potential (E(p)) upon microelectrode impalement is a more accurate estimate of the true macrophage membrane potential than the sustained potential (E(s)).
- Slow hyperpolarizations observed after impalement represent transient repolarizations back to the actual membrane potential.
- Analysis of the initial impalement transient is a valuable technique for estimating the membrane potential of small, isolated cells.