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PATCH CLAMP TECHNIQUES

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[1] Patch Clamp Techniques: An Overview By M I C H A E L

C A H A L A N a n d ERWIN N E H E R

Historical Introduction Patch-recording techniques originated from an effort to record currents through individual ion channels in biological membranes. By the early 1970s it had become clear that discrete molecular entities--integral membrane proteins-- underlie the electrical signaling mechanisms of nerve and muscle. The selective actions of certain toxins, proteases, and proteinmodifying agents had indicated that sodium and potassium channels constituted separate macromolecules. 1-3 Studies in artificial lipid membranes had shown that certain proteins isolated from bacteria 4 and some antibiotic polypeptides s were able to induce steplike changes in membrane conductance, which were attributed to the opening and closing of individual pore- or channellike structures. These discrete conductance changes were of the same order of magnitude as single-channel conductances inferred from analysis of current fluctuations in the neuromuscular junction and in nodes of Ranvier. 6-s Thus, it was tempting to look for similar discrete current changes in biological preparations. Bilayer studies had shown that electronic components available at that time were capable of handling such small signals. On the other hand, background noise in all standard voltage-clamp arrangements was higher by one to two orders of magnitude than the signals to be measured. Thus, it seemed straightforward to attempt to isolate a small area of membrane surface (a "patch") for localized electrical measurement by placing a measuring glass micropipette onto the surface of a voltageclamped cell. Such arrangements had already been used for focal stimulation 9 or for measurement of local current density? °-~2 Simple consideraJ. W. Moore and T. Narahashi, Fed. Proc. 26, 1655 (1967). 2 B. Hille, J. Gen. Physiol. 50, 1287 (1967). 3 C. M. Armstrong, F. Bezanilla, and E. Rojas, J. Gen. Physiol. 62, 377 (1973). 4 R. C. Bean, W. C. Shepherd, H. Chan, and J. Eichner, J. Gen. Physiol. 53, 741 (1969). 5 S. B. Hladky and D. A. Haydon, Nature (London_)225, 451 (1970). 6 B. Katz and R. Miledi, J. Physiol. (London) 224, 665 (1972). 7 C. R. Anderson and C. F. Stevens, J. Physiol. (London) 235, 651 (1973). s F. Conti, B. Hille, and W. Nonner, J. Physiol. (London) 353, 199 (1984). 9 A. F. Huxley and R. E. Taylor, J. Physiol. (London) 144, 426 (1958). to A. Stricldaolm, J. Gen. Physiol. 44, 1073 (1961). LK. Frank and L. Tauc, in "The Cellular Function of Membrane Transport" (J. Hoffman, ed.), p. 113. Prentice-Hall, Englewood Cliffs, New Jersey, 1963. 12 E. Neher and H. D. Lux, PfluegersArch. 311, 272 (1969).

METHODSIN ENZYMOLOGY,VOL 207

~ t © 1992by AcademicPre~ Inc. Allt~ghtsof reproductionin any formreserved.

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tions of background noise led to the conclusion that such pipettes should allow resolution of picoampere-sized currents, such as acetylcholine-induced responses, whenever the "seal-resistance" - - the resistance between the interior of the pipette and the bath--could be increased severalfold above the pipette internal resistance a n d be made to exceed a final value of 50-100 M ~ (for marginal resolution). Unfortunately, the initial experiments showed that it was very dilficult to obtain satisfactory seals. In spite of systematic investigation using enzymes to clean cell surfaces, using a variety of pipette sizes and shapes, and using pipette glass with varying surface charge and hydropathy, it seemed impossible to improve the seal resistance beyond 100 M~. Nevertheless, optimizing enzymatic cleaning procedures and pipette geometries allowed single acetylcholine-induced currents to be resolved.13 The major drawbacks of measurements with low-resistance seals (between 1976 and 1980) included limited resolution, frequent occurrence of partial single-channel events owing to channels localized in the pipette rim area, and large leakage currents associated with excessive noise whenever small voltage differences occurred between pipette interior and bath. All these problems were eliminated or greatly improved when it was found 14,15 that the application of slight suction within a freshly prepared pipette readily induced the phenomenon now called gigaseal formation. This is a sudden transition of seal resistance from several tens of megohms to several gigaohms. The physical basis of the gigaseal is still not quite clear, but it certainly increased resolution by an order of magnitude, decreased the variability of channel step sizes, and allowed potentials to be applied across the seal for local voltage stimulation. Furthermore, it was recognized that the gigaseal provided mechanical stability, so that patches could be "excised" from the parent cell and studied in isolation.16 Ironically, breaking patches paved the way to another present-day application of the patchclamp, tight-seal whole-cell recording. Breaking the patch (without the loss of the seal) provides electrical continuity between the patch pipette and the cell interior, thus leading to a configuration similar to conventional microelectrode impalement. It turned out that this form of impalement was much gentler than conventional microelectrode methods, and that it was tolerated by cells as small as a few micrometers in diameter. In small cells, the membrane impedance can be orders of magnitude higher than the 13E. Neherand B, Sakmann,Nature (London) 260, 779 (1976). 14F. J. Sigworthand E. Neher,Nature (London) 287, 447 (1980). 1~O. P. Hamill, A. Marty,E. Neher,B. Sakmann,and F. J. Sigworth,PfluegersArch. 391, 85 (1981). 16R. Horn and J. Patlak, Proc. Natl. Acad. Sci. U.S.A. 77, 6930 (1980).

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internal resistance of the measuring pipette. This is quite different from the case of conventional impalements, in which pipette resistance and cell impedance are usually of the same order of magnitude, requiring feedback circuitry for voltage clamping. Procedures for obtaining the basic patch clamp configurations were well established by late 1980, as described by Hamill et a l ) s Some of the historical details of the development have been considered by Sigworth. 17 Experience from laboratories up to 1983 has been summarized in SingleC h a n n e l Recording, edited by Sakmann and Neher) s Whereas the electrical aspects of patch clamp measurement consolidated within a year or two, it took somewhat longer to appreciate fully the biochemical implications of the technique. We now know that ion channels are gated and modulated not only by voltage and external ligands, but also by second messengers, regulatory proteins, and by phosphorylation. Many of these regulatory molecules are definitely lost when a patch is excised; even in the whole-cell recording mode, mobile intracellular ions may exchange with those in the pipette within seconds, t9.2° and second messengers may be lost within minutes. This feature may be utilized to investigate the influence of such regulators. For this purpose, techniques have been developed to perfuse the pipette interior (see [ 10] in this volume). On the other hand, "washout" of regulators may prevent prolonged study of physiological functions in small cells. For this reason, efforts have been made to find ways to permeabilize patches for small ions selectively, in order to provide electrical access to the cell without perturbing the cellular biochemistry. These techniques are known as slow whole-cell,2~ perforated-patch, or the nystatin method 22 (see also [8] in this volume). Patch Configurations Here, we provide an introduction to the variety of recording configurations that are possible following formation of a gigaseal. Four of them are essentially as described previously by HamiU et al. t5 and summarized in the flowchart of Fig. 1. In addition, perforated-patch recording and other minor variants are summarized briefly. L7F. J. Sigworth,Fed. Proc. 45, 2673 0986). 18B. Sakmann and E. Neher, eds., "Single-ChannelRecording."Plenum, New York and London, 1983. ~9A. Marryand E. Neher, in "Single-ChannelRecording"(B. Sakmannand E. Neher,eds.). Plenum, New York and London, 1983. 2oM. Puschand E. Neher,Pfluegers Arch. 411, 204 (1988). 21 i . Lindau and J. i . Fernandez, Nature (London) 319, 150 (1986).

22R. Horn and A. Marty,J. Gen. Physiol. 92, 145 (1988).

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ELECTROPHYSIOLOGICAL TECHNIQUES cell-attached

[ 1]

inside-out

-5O

perforated- patch

whole - cell

outside-out

FIo. 1. Schematic diagram of patch clamp configurations. The numbers indicate potenrials in millivolts and have been chosen so that a membrane potential o f - 50 mV is applied in all the configurations, assuming a cell resting potential o f - 70 mV. A pipette is moved toward the cell surface (upper left) to form a gigaseal. From the cell-attached configuration, one can attain (l) the inside-out configuration by withdrawing the pipette; (2) the whole-cell configuration by rupturing the patch using a pulse of suction or voltage; and (3) the perforated patch by including nystatin in the pipette filling solution. From the whole-cell configuration, one can attain the outside-out patch configuration by withdrawing the pipette (see text for details).

Seal Formation As the pipette is advanced toward the cell, the pipette resistance is monitored by repetitively applying a small (usually < 10 mV) voltage step to the pipette. Typically, fire-polished glass pipettes with resistances of the order of 1 - 10 Mfl are employed, corresponding to tip diameters of the order of 1/zm. Signals that are applied, and that can be observed during "sealing," are illustrated in Fig. 2. When the pipette touches the cell, the resistance increases. Application of suction to the interior of the pipette (typically 10-20 cm H20) draws a small portion of cell membrane into the pipette, and, if all is "right," seal formation is visualized as a sudden increase in resistance such that virtually no current can pass between pipette and bath electrodes. When the resistance is measured carefully at high gain, seals of over 10 ~° fl are common. Several factors promote seal formation, including Millipore-filtered solutions, positive pressure within the pipette during the approach to the cell, and a clean cell surface. Pipette glass and the composition of solutions also play a role in determining

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PATCH CLAMP TECHNIQUES

Seal Formation

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Cell-attached and Break-in

-I 33 mV

40 mV I

500 pA

100 pA 1 ms

I

10 ms

L

....

/

-[ .......

................. L. . . . . . . .

FIG. 2. Typical electrical signals observed during sealing and break-in. In the left-hand column (sealing), a 5-mV negative pulse is applied (top trace). The resulting currents are shown in the traces below. The top current trace was obtained before the pipette touched the cell. The amplitude of the current pulse indicates a pipette resistance of 4.4 Mt~ (Ohm's law!). The center trace indicates a somewhat increased pipette resistance (smaller signal) after the pipette touched the cell. The bottom trace was recorded a few seconds after suction had been applied. The current signal virtually disappeared because a gigas~ had formed. The only signal visible at this resolution is a small transient of capacitive charging current due to the stray capacitance of the pipette. In the fight-hand column, the voltage pulse was increased to 50 mV; current sensitivity was also increased. The capacitive transients on the current trace (see above) would be expected to be sizable under these conditions; however, they have been almost eliminated by fast capacitance neutralization. Subsequently, a pulse of suction was given, resulting in the third trace. Two changes can be seen when compared to the second trace: (1) an increase in noise and (2) large capacitive transients which have, in fact, been truncated. Both changes indicate patch rupture. The transients are due to the capacitance of the cell. The increased noise is due to the ionic conductances and the capacitance oftbe cell. In the bottom trace, the cell capacitance has been neutralized by electronic capacitance compensation. At this stage, a series of voltage pulses can be applied, as shown in Fig. 3.

success in forming seals. When conditions are favorable, seal formation occurs in nearly 100% of trials.

Cell-AttachedPatch Recording Currents through ion channels trapped within the pipette orifice can be recorded with subpicoampere resolution following seal formation. The overall sensitivity of the measurement depends on a variety of factors, including electronic noise, pipette capacitance, noise associated with the holder, and "pickup" of line frequency or computer-associated noise of

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higher frequency. Between 0 and 300 Hz, a background noise value of the order of 0.1 pA root mean square (rms) can be achieved using a 50-G~ feedback resistor or capacitative feedback for current measurement, enabling detection of currents through single channels. If the cell is electrically active, action potentials will be detected as a capacitative current across the membrane patch. In this recording configuration, the cell remains intact, although seal formation involves some deformation as membrane is sucked into the pipette. The potential across the membrane is equal to the membrane potential of the cell minus the pipette potential. Applying a negative potential to the pipette depolarizes the patch of membrane. If channels within the patch are active, positive current flowing outward across the membrane goes into the pipette, and negative current flowing inward across the membrane goes out of the pipette. Because the seal will not permit leakage of bath constituents into the pipette, the extracellular surface of the patch is exposed only to the pipette solution; bath solution changes can be made without altering the solution exposed to the patch.

Inside-out Patch An isolated patch of membrane can be torn off the cell simply by withdrawing the pipette after seal formation. Normally, the cell survives this insult. If the patch is excised from the cell-attached recording configuration, the resulting patch is "inside-out," with the cytoplasmic membrane surface facing the bath solution. Again, from the standpoint of the membrane, positive current flowing outward across the membrane goes into the pipette. However, in this case the membrane potential of the patch is the bath potential minus the pipette potential. Thus, to achieve a membrane potential o f - 50 mV, for example, a pipette potential o f + 50 mV must be applied. Single-channel resolution can be improved by lifting the patch near the surface of the bath solution in order to reduce the capacitance of the pipette. One problem which can occur is the formation of a "vesicle" if the patch of membrane seals over, enclosing a small volume of bathing solution. Sometimes this can be corrected by brief exposure of the pipette tip to air. Vesicle formation is inhibited by bath solutions containing low amounts of calcium. Excised patches do not consist solely of the membrane. Cytoskeletal elements and membranous organelles may also be present, as demonstrated vividly by a recent description of light-activated photoreceptor current in patches from retinal rods. 2a In this case the entire cytoplasmic transduction pathway including rhodopsin, transducin, phos23 E. A. Ertel, Proc. Natl. Acad. Sci. U.S.A. 87, 4226 (1990).

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25 mvl~ S ms

500 pA [ 5 ms Fie. 3. Series of superimposed current records from whole-cell recording in response to depolarizing stimuli to -20, - 1 0 , 0, and + 10 mV from a holding potential o f - 7 0 mV, using a bovine chromalfin cell at room temperature. The cell capacitance of 5 pF has been compensated (series resistance 7.2 M~).

phodiesterase, and cyclic GMP was present in the excised patch, in addition to the sodium channel present in the membrane.

Whole-Cell Recording Following initial seal formation, it is also possible to rupture deliberately the patch of membrane trapped within the pipette by application of strong suction or a voltage pulse of several hundred millivolts. Patch rupture is detected as a sudden increase in capacitative current in response to a test potential step, as the membrane capacitance of the cell is "seen" through the series resistance of the pipette. This "break-in" results in electrical and diffusional continuity between the pipette and the cytoplasm, usually without altering the seal resistance between glass and membrane. If the access resistance between the pipette electrode and the cytoplasm is much lower than the membrane resistance of the cell, as is usually the case in small cells, the membrane potential of the cell is thereby voltage clamped to the pipette potential, with polarity opposite that of the insideout patch; the membrane potential of the cell will be equal to the pipette potential minus the bath potential. The ensemble activity of ion channels can then be detected by monitoring the current. In the case of whole-cell recording, positive, outward membrane current flows out of the pipette. In the example shown in Fig. 3, inward sodium currents and outward potassium currents in response to a family of

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voltage-clamp steps are illustrated. As pointed out above, whole-cell recording permits diffusional exchange of pipette contents with cytoplasmic constituents; ionic contents can be exchanged within seconds. From the point of view of lacing able to control both the electrical potential and cytoplasmic milieu, one major factor is maintaining good access between pipette and cytoplasm. If membrane currents are large (exceeding the nanoampere range), compensation for series resistance can help control the membrane potential at the desired level by electronic feedback. However, geometrical factors must also be considered in achieving adequate voltage control. These considerations are discussed in [5] in this volume.

Outside-out Patch If the pipette is withdrawn from the cell during whole-cell recording, a tether of membrane is drawn away from the cell which normally breaks off and reseals on the pipette as an outside-out patch. Voltage and polarity conventions are identical to whole-cell recording, but the membrane surface area is greatly reduced, allowing subpicoampere resolution of current. In a variant of this technique, the nucleus of the cell is carded along during patch excision resulting in an outside-out "macropatch. ''24

Perforated-Patch Recording An unavoidable consequence of whole-cell recording is the loss of cytoplasmic ions, nucleotides, and other diffusible constituents into the pipette. Several techniques have been invented to achieve electrical continuity between pipette and cytoplasm while minimizing dialysis. Poreforming antibiotic molecules such as nystatin can be added to the pipette solution following seal formation, or backfilled at some distance from the tip to allow for normal seal formation. Nystatin will then spontaneously form conducting channels selective for monovalent ions, and thereby lower the electrical resistance of the patch to the point where the cell can be voltage clamped and current measured through the permeabilized patch of membrane beneath the pipette. This has the great advantage that secondmessenger mechanisms within the cell can remain intact during the recording. Electronic Enhancements The design of modern, commercially available patch-clamp amplifiers incorporates low-noise current-measuring circuitry, voltage summing to deliver holding, offset, and applied command potentials to the pipette, 24 p. Aschcr, Soc. Neurosci. Abstr. 16, 619 (1990).

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capacity current subtraction incorporating at least two time constants, and electronic feedback compensation for series resistance. Low-pass filtering, internal pulse generation, voltage outputs to indicate gain and filter settings, and tone generators for auditory monitoring of seal resistances are additional features often provided. Some designs incorporate headstages with switchable feedback resistors to allow optimal signal-to-noise ratios for high-gain recording, while allowing large currents to be monitored without saturation at low gain. Other designs incorporate capacitative feedback to monitor current with great sensitivity. The voltage clamp can be interfaced to a digital computer enabling the programming of sophisticated stimulation and data acquisition protocols. In some designs, a computer can control the status of the amplifier directly, providing a more direct linkage between software and hardware. Applications

Single-Channel Recording Perhaps the most startling and inherently pleasing outcome of patch clamp methodology is simply the ability to monitor, in real time, the dynamics of a single ion channel within a membrane patch. Single-channel currents through most ion channel types have already been detected. Currents through single ion channels appear when conformational changes in the channel protein gate the flow of thousands of ions per millisecond. The detailed kinetic study of single-channel currents has provided unique insights into the mechanisms of ion-channel gating. A further example of the power of combining molecular and electrophysiological approaches is the investigation of the molecular genetics and function of ion channels. Patch clamp techniques are ideal for detailed analysis of channel kinetic states. The analysis of site-directed mutations with patch clamp techniques using Xenopus oocytes and transfected mammalian cells is a focus of much current research on the structure and function of ion channels. These topics are reviewed elsewhere in this volume.

Expanded Scope of Electrophysiology Initially, patch damp recordings were performed mainly on acutely isolated and enzymatically cleaned preparations. Soon it became dear, however, that the method was ideally suited for basically all cell culture preparations. At first, the major focus was to characterize ion channels at the single-channel level in order to determine unit conductance values and to investigate biophysical models for channel gating. However, the ability

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to investigate ion channels in a wide variety of cell types has vastly expanded the scope of modern electrophysiology. Quantitative voltage clamp measurements are no longer limited to a few preparations which, by virtue of size or special geometry, were especially favorable for voltage control by electronic feedback. Among the isolated or cultured cell preparations that have been investigated are a wide variety of neurons; cells of sensory transduction; glial cells; muscle cells including skeletal, cardiac, and smooth muscle types; endothelial and epithelial cells including those in the vasculature, the respiratory system, and the kidney; secretory cells including pancreatic acinar cells, lacrimal gland cells, and juxtaglomerular cells; hepatocytes; pancreatic fl cells; keratinocytes, osteoblasts, and osteoelasts; and cells of hematopoietic origin including erythrocytes, lymphocytes, macrophages, mast cells, neutrophils, and blood platelets. Methods have also been adapted to extend measurements to o o c y t e s , 25 plant protoplasts, 26 yeast, 27 and bacteria. 27 In addition to measurements on isolated cells, procedures have been described to obtain tight seals on defined cells in brain slices,2s a technique which allows functional connections in the central nervous system (CNS) to be studied at unsurpassed resolution. Recently, explorations of ion channels in subcellular organelles have begun, including studies on mitochondria,29 the nucleus,3° and endoplasmic reticulum? 1 Techniques to record from liposomes or to form phospholipid bilayers on patch clamp pipettes have also been described,a2,33

Discovery of Novel Ion Channels The patch clamp has also expanded the catalog of recognized ion channel subtypes. It is now clear from patch clamp and from molecular biological approaches that voltage-dependent sodium, calcium, and potassium channels constitute a large superfamily composed of a wide variety of subtypes. A second major superfamily includes the transmitter-gated channel/receptors. In addition, it is now clear that distinct channel species 25 C. Methfessel, V. Witzemann, T. Takahashi, M. Mishina, S. Numa, and B. Sakmann, Pfluegers Arch. 407, 577 (1986). 26 R. Hedrich and J. Schroeder, Annu. Rev. Plant Physiol. 40, 539 (1989). 27 y. Saimi, B. Martinac, M. R. Culbertson, J. Adler, and C. Kung, Cold Spring Harbor Syrup. Quant. Biol. 53, 667 (1988). 2s A. Konnerth, Trends Neurosci. 13, 321 (1990). 29 M. C. Sorgato, B. U. Keller, and W. Stuhmer, Nature (London) 330, 498 (1987). 3oM. Mazzanti, L. J. DeFelice, J. Cohen, and H. Malter, Nature (London) 343, 764 (1990). 3~A. Schmid, M. Dehlinger-Kremer, I. Sehultz, and H. Gogelein, Nature (London) 346, 374 (1990). 32 D. W. Tank, C. Miller, and W. W. Webh, Proc. Natl. Acad. Sci. U.S.A. 79, 7749 0982). 33 R. Coronado and R. Latorre, Biophys. J. 43, 231 (1983).

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that have not as yet been characterized at the molecular level are regulated by nueleotides, intracellular calcium and sodium ions, and by GTP-binding proteins. As a result, instead of the handful of channels recognized before the mid-1970s, hundreds of distinct channel subtypes that are regulated by a variety of mechanisms have now been described.

Capacitance Measurements Current through a membrane consists of an ionic as well as a capacitafive component. Because the fundamental structure of biological membranes consists of a lipid bilayer in which intrinsic membrane proteins are immersed, a value of approximately 1 #F/cm 2 is considered to represent the value of membrane capacitance. One picofarad of membrane capacitance represents approximately 100 #m 2 of membrane surface area. Because current measurement is extremely sensitive during patch recording, it is possible to measure the membrane area with great accuracy by monitoring membrane capacitance. This can be done by applying voltage steps and measuring the capacitative current response, in which case membrane capacitance is proportional to the integral of the charging transient. Alternatively, a lock-in amplifier which measures current in and out of phase with a sinusoidal applied potential provides resolution of capacitance to approximately 10 fF. The sensitivity of this measurement is such that the fusion of single secretory vesicles in mast cells has been measured as a step increase in membrane capacitance. ~,3s Thus, patch recording has enabled not only single channels to be resolved, but also single exocytotic events. In whole-cell recording, capacitance provides a real-time measure of secretion which has been applied to several cell types.

Cell-Signaling Mechanisms Ion channels not only mediate electrical excitability in the nervous system and in the heart; they also appear to play important functional roles in the behavior of most cell types. One important factor in considering cell-signaling mechanisms is that the activity of ion channels can directly or indirectly affect the concentration of an important second messenger, namely, calcium ions. In most excitable cells, voltage-dependent calcium channels not only help to shape the action potential, but they gate the entry of calcium ions which in turn can activate kinases, contractile proteins, or ion channels. In a variety of electrically inexcitable cells, voltage-indepen34 E. Neher and A. Marty, Proc. Natl. Acad. Sci. U.S.A. 79, 6712 (1982). 35 M. Lindau and E. Neher, PfluegersArch. 411, 137 (1988).

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dent calcium channels can be activated following the binding o f a ligand to receptors on the cell surface. Furthermore, both receptor-linked GTPbinding proteins and second-messenger systems within the cytosol can affect the activity o f ion channels. Thus, complex regulatory pathways linking surface receptors, the metabolism of the cell and ion channels can result in long-lasting changes in the behavior of the cell. The patch clamp technique provides the experimental means for merging the tools of modern molecular and cellular biology with those of electrophysiology. Using the various recording configurations, it is possible to dissect the mechanisms of channel modulation. In cell-attached recording, modulation of channel activity in response to bath-applied agonist generally indicates a second-messenger mechanism. Candidate messengers can be tested directly on excised patches or in whole-cell recording. Perforated patch recording maintains the integrity of second-messenger systems while enabling the overall activity of ion channels in the cell to be evaluated following receptor stimulation. Current research on signaling pathways seeks to establish the functionally meaningful mechanisms through selective activation or inhibition of a portion of the pathway. The variety of patch clamp configurations, combined with single-channel resolution, provides a powerful experimental approach from the molecular level, in which channel genes are altered and expressed, to the cellular level, in which posttranslational signaling mechanisms are elucidated, to the systems level, in which cellular interactions in intact or slice preparations are revealed.

[2] C o n s t r u c t i n g

A Patch

Clamp

Setup

By RICHARD A. LEVIS and JAMES L. RAE

Basic C o m p o n e n t s in P a t c h Clamp Setup Different investigators have chosen to construct their patch clamp setups in very different ways, and it is clear that there is no one "best" way to configure the apparatus for patch clamping. There are, however, many features that good setups have in common, and there are some basic principles that one should consider when configuring and purchasing patch clamp hardware) ,2 In this chapter we discuss these principles, describe t j. L. Rae and R. A. Levis,Mol. Physiol. 6, 115 (1984). 2j. L. Rae, R. A. Levis,and R. S. Elsenberg,in "Ion Channels"(T. Narahashi, ed.), p. 283. Plenum, New York and London, 1988. METHODS IN ENZYMOLOGY, VOL. 207

Copyright © 1992 by Ac~lemic Press, Inc. All fights of repr~uction in any form reserved.

Patch clamp techniques: an overview.

[ 1] PATCH CLAMP TECHNIQUES 3 [1] Patch Clamp Techniques: An Overview By M I C H A E L C A H A L A N a n d ERWIN N E H E R Historical Introductio...
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