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Intracellular recordings of action potentials by an extracellular nanoscale field-effect transistor.

Duan X, Gao R, Xie P, Cohen-Karni T, Qing Q, Choe HS, Tian B, Jiang X, Lieber CM - Nat Nanotechnol (2011)

Bottom Line: Field-effect transistors (FETs) can also record electric potentials inside cells, and because their performance does not depend on impedance, they can be made much smaller than micropipettes and microelectrodes.Simulations show that the bandwidth of this branched intracellular nanotube FET (BIT-FET) is high enough for it to record fast action potentials even when the nanotube diameter is decreased to 3 nm, a length scale well below that accessible with other methods.We also show that multiple BIT-FETs can record multiplexed intracellular signals from both single cells and networks of cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts 02138, USA.

ABSTRACT
The ability to make electrical measurements inside cells has led to many important advances in electrophysiology. The patch clamp technique, in which a glass micropipette filled with electrolyte is inserted into a cell, offers both high signal-to-noise ratio and temporal resolution. Ideally, the micropipette should be as small as possible to increase the spatial resolution and reduce the invasiveness of the measurement, but the overall performance of the technique depends on the impedance of the interface between the micropipette and the cell interior, which limits how small the micropipette can be. Techniques that involve inserting metal or carbon microelectrodes into cells are subject to similar constraints. Field-effect transistors (FETs) can also record electric potentials inside cells, and because their performance does not depend on impedance, they can be made much smaller than micropipettes and microelectrodes. Moreover, FET arrays are better suited for multiplexed measurements. Previously, we have demonstrated FET-based intracellular recording with kinked nanowire structures, but the kink configuration and device design places limits on the probe size and the potential for multiplexing. Here, we report a new approach in which a SiO2 nanotube is synthetically integrated on top of a nanoscale FET. This nanotube penetrates the cell membrane, bringing the cell cytosol into contact with the FET, which is then able to record the intracellular transmembrane potential. Simulations show that the bandwidth of this branched intracellular nanotube FET (BIT-FET) is high enough for it to record fast action potentials even when the nanotube diameter is decreased to 3 nm, a length scale well below that accessible with other methods. Studies of cardiomyocyte cells demonstrate that when phospholipid-modified BIT-FETs are brought close to cells, the nanotubes can spontaneously penetrate the cell membrane to allow the full-amplitude intracellular action potential to be recorded, thus showing that a stable and tight seal forms between the nanotube and cell membrane. We also show that multiple BIT-FETs can record multiplexed intracellular signals from both single cells and networks of cells.

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Intracellular action potential recordinga, a representative trace reflects the transition from extra- to intra- cellular recording. b, magnified trace of the part in the black dashed rectangle in a. c, magnified trace of the peak in the blue dashed rectangle in b. The stars in b and c mark the position of extracellular spikes. d, magnified trace of the peak in red dashed rectangle in a. e, the trace corresponding to the second entry of the nanotube around the same position on the cell. The potential was calibrated using the sensitivity values measured on phospholipid-modified devices by quasi-static Vwg measurement (e.g. blue trace in Fig. 2c) and pulsed Vwg measurement with 0.1 ms pulse rise/fall time (same for Fig. 4). The sensitivity obtained from these two measurements is same as discussed before.
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Figure 3: Intracellular action potential recordinga, a representative trace reflects the transition from extra- to intra- cellular recording. b, magnified trace of the part in the black dashed rectangle in a. c, magnified trace of the peak in the blue dashed rectangle in b. The stars in b and c mark the position of extracellular spikes. d, magnified trace of the peak in red dashed rectangle in a. e, the trace corresponding to the second entry of the nanotube around the same position on the cell. The potential was calibrated using the sensitivity values measured on phospholipid-modified devices by quasi-static Vwg measurement (e.g. blue trace in Fig. 2c) and pulsed Vwg measurement with 0.1 ms pulse rise/fall time (same for Fig. 4). The sensitivity obtained from these two measurements is same as discussed before.

Mentions: We investigated the capability of the BIT-FET to record intracellular signals using spontaneously beating embryonic chicken cardiomyocyte cells, which were cultured on thin pieces of polydimethylsiloxane (PDMS) as described previously16. After modifying the devices with phospholipids10 to facilitate the internalization of nanotubes into cells, the PDMS/cell sheet was manipulated to put a cell into gentle contact with the nanotube of a BIT-FET under standard electrophysiology microscope (see Supplementary Methods). Approximately 45s after gentle contact was made and in the absence of applied force to the cell substrate, the recorded data showed a dramatic change (Fig. 3a). Before the transition, the signal exhibits a relatively flat baseline with small biphasic peaks (5~8 mV amplitude; ~1 ms duration) with ca. 1 Hz frequency (e.g., Figs. 3b, c). These peaks are coincident with cell beating and consistent with extracellular recording reported previously16. Then the baseline shifts ca. -35 mV and new peaks with 75-100 mV amplitude and ~200 ms duration are observed (Fig. 3a). The recorded conductance data yields inverted peaks for the p-type SiNW FETs used here, although the calibrated potentials are consistent with standard peak polarity and shape of intracellular action potentials. These peaks (e.g., Fig. 3d) have the shape and features characteristic of an intracellular action potential of cardiomyocyte cells10,19,20, including fast depolarization at the beginning of the peak, plateau region, fast repolarization, and hyperpolarization and return to baseline. The signal transition from extra- to intracellular indicated the penetration of the cell by the nanotube. The baseline shift is similar with that measured recently using kinked-nanowire probes10, but smaller than the standard resting potential for cardiomyocytes19,20. Our reproducible and stable recording of full-amplitude action potentials, which is a central result of our work, suggests that this baseline difference is not due to poor sealing during nanotube internalization. We propose that the discrepancy in resting potentials here could be attributed to a stronger suspension effect introduced by the intracellular polyelectrolytes at the junction21,22 due to an order of magnitude smaller size of SiO2 nanotube opening than a typical patch clamp pipette, although more detailed studies will be required to quantitatively understand the origin of this effect. Although the nanotube diameter routinely used in our intracellular recording studies, 50 nm inner diameter and 55 nm tip outer diameter, is larger than the smallest achievable for BIT-FETs (Fig. 2f), it is still much smaller than the size of typical glass micropipettes1,2 and metal microelectrodes3,4,7 used for intracellular studies.


Intracellular recordings of action potentials by an extracellular nanoscale field-effect transistor.

Duan X, Gao R, Xie P, Cohen-Karni T, Qing Q, Choe HS, Tian B, Jiang X, Lieber CM - Nat Nanotechnol (2011)

Intracellular action potential recordinga, a representative trace reflects the transition from extra- to intra- cellular recording. b, magnified trace of the part in the black dashed rectangle in a. c, magnified trace of the peak in the blue dashed rectangle in b. The stars in b and c mark the position of extracellular spikes. d, magnified trace of the peak in red dashed rectangle in a. e, the trace corresponding to the second entry of the nanotube around the same position on the cell. The potential was calibrated using the sensitivity values measured on phospholipid-modified devices by quasi-static Vwg measurement (e.g. blue trace in Fig. 2c) and pulsed Vwg measurement with 0.1 ms pulse rise/fall time (same for Fig. 4). The sensitivity obtained from these two measurements is same as discussed before.
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Related In: Results  -  Collection

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Figure 3: Intracellular action potential recordinga, a representative trace reflects the transition from extra- to intra- cellular recording. b, magnified trace of the part in the black dashed rectangle in a. c, magnified trace of the peak in the blue dashed rectangle in b. The stars in b and c mark the position of extracellular spikes. d, magnified trace of the peak in red dashed rectangle in a. e, the trace corresponding to the second entry of the nanotube around the same position on the cell. The potential was calibrated using the sensitivity values measured on phospholipid-modified devices by quasi-static Vwg measurement (e.g. blue trace in Fig. 2c) and pulsed Vwg measurement with 0.1 ms pulse rise/fall time (same for Fig. 4). The sensitivity obtained from these two measurements is same as discussed before.
Mentions: We investigated the capability of the BIT-FET to record intracellular signals using spontaneously beating embryonic chicken cardiomyocyte cells, which were cultured on thin pieces of polydimethylsiloxane (PDMS) as described previously16. After modifying the devices with phospholipids10 to facilitate the internalization of nanotubes into cells, the PDMS/cell sheet was manipulated to put a cell into gentle contact with the nanotube of a BIT-FET under standard electrophysiology microscope (see Supplementary Methods). Approximately 45s after gentle contact was made and in the absence of applied force to the cell substrate, the recorded data showed a dramatic change (Fig. 3a). Before the transition, the signal exhibits a relatively flat baseline with small biphasic peaks (5~8 mV amplitude; ~1 ms duration) with ca. 1 Hz frequency (e.g., Figs. 3b, c). These peaks are coincident with cell beating and consistent with extracellular recording reported previously16. Then the baseline shifts ca. -35 mV and new peaks with 75-100 mV amplitude and ~200 ms duration are observed (Fig. 3a). The recorded conductance data yields inverted peaks for the p-type SiNW FETs used here, although the calibrated potentials are consistent with standard peak polarity and shape of intracellular action potentials. These peaks (e.g., Fig. 3d) have the shape and features characteristic of an intracellular action potential of cardiomyocyte cells10,19,20, including fast depolarization at the beginning of the peak, plateau region, fast repolarization, and hyperpolarization and return to baseline. The signal transition from extra- to intracellular indicated the penetration of the cell by the nanotube. The baseline shift is similar with that measured recently using kinked-nanowire probes10, but smaller than the standard resting potential for cardiomyocytes19,20. Our reproducible and stable recording of full-amplitude action potentials, which is a central result of our work, suggests that this baseline difference is not due to poor sealing during nanotube internalization. We propose that the discrepancy in resting potentials here could be attributed to a stronger suspension effect introduced by the intracellular polyelectrolytes at the junction21,22 due to an order of magnitude smaller size of SiO2 nanotube opening than a typical patch clamp pipette, although more detailed studies will be required to quantitatively understand the origin of this effect. Although the nanotube diameter routinely used in our intracellular recording studies, 50 nm inner diameter and 55 nm tip outer diameter, is larger than the smallest achievable for BIT-FETs (Fig. 2f), it is still much smaller than the size of typical glass micropipettes1,2 and metal microelectrodes3,4,7 used for intracellular studies.

Bottom Line: Field-effect transistors (FETs) can also record electric potentials inside cells, and because their performance does not depend on impedance, they can be made much smaller than micropipettes and microelectrodes.Simulations show that the bandwidth of this branched intracellular nanotube FET (BIT-FET) is high enough for it to record fast action potentials even when the nanotube diameter is decreased to 3 nm, a length scale well below that accessible with other methods.We also show that multiple BIT-FETs can record multiplexed intracellular signals from both single cells and networks of cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts 02138, USA.

ABSTRACT
The ability to make electrical measurements inside cells has led to many important advances in electrophysiology. The patch clamp technique, in which a glass micropipette filled with electrolyte is inserted into a cell, offers both high signal-to-noise ratio and temporal resolution. Ideally, the micropipette should be as small as possible to increase the spatial resolution and reduce the invasiveness of the measurement, but the overall performance of the technique depends on the impedance of the interface between the micropipette and the cell interior, which limits how small the micropipette can be. Techniques that involve inserting metal or carbon microelectrodes into cells are subject to similar constraints. Field-effect transistors (FETs) can also record electric potentials inside cells, and because their performance does not depend on impedance, they can be made much smaller than micropipettes and microelectrodes. Moreover, FET arrays are better suited for multiplexed measurements. Previously, we have demonstrated FET-based intracellular recording with kinked nanowire structures, but the kink configuration and device design places limits on the probe size and the potential for multiplexing. Here, we report a new approach in which a SiO2 nanotube is synthetically integrated on top of a nanoscale FET. This nanotube penetrates the cell membrane, bringing the cell cytosol into contact with the FET, which is then able to record the intracellular transmembrane potential. Simulations show that the bandwidth of this branched intracellular nanotube FET (BIT-FET) is high enough for it to record fast action potentials even when the nanotube diameter is decreased to 3 nm, a length scale well below that accessible with other methods. Studies of cardiomyocyte cells demonstrate that when phospholipid-modified BIT-FETs are brought close to cells, the nanotubes can spontaneously penetrate the cell membrane to allow the full-amplitude intracellular action potential to be recorded, thus showing that a stable and tight seal forms between the nanotube and cell membrane. We also show that multiple BIT-FETs can record multiplexed intracellular signals from both single cells and networks of cells.

Show MeSH