Displacement Photocurrents in Pigment-Containing Biomembranes

14 Displacement Photocurrents in

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Pigment-Containing Biomembranes: Artificial and Natural Systems F E L I X T. H O N G Department

of Physiology,

Detroit, M I 48201

Wayne

State

University,

School of Medicine,

A review of selected literature concerned with displacement photoelectric currents in pigment-containing biomembranes is presented. From model system studies, it appears that the displacement photocurrent is a manifestation of a chemical capacitance rather than the ordinary membrane capacitance. Analysis indicates that observed relaxation of the displacement photoelectric responses is influenced critically by the particular method of measurement. While the tunable voltage clamp (quasipotentiostatic) method provides relevant kinetic data, the open-circuit (galvanostatic) measurement preserves little kinetic information. Two different molecular models (oriented dipole vs. interfacial charge transfer) are discussed in light of current insight and with special reference to visual membranes and purple membranes of Halobacterium halobium. "p\isplacement (capacitative) photocurrents have been found i n a wide variety of pigment-containing biomembranes. The best known example is the early receptor potential ( E R P ) , which was discovered i n 1964 by Brown and Murakami i n the frog retina (1,2,3). The most salient feature of the E R P that distinguishes it from other bioelectric phenomena is the apparent lack of a latency (4). It is also remarkable for its resistance to anoxia and to drastically altered ionic environments (1,5,6,7,8). The E R P has two components (see Figure 1 ) : a corneapositive and temperature-insensitive R l component and a cornea-negative R2 component, which is inhibited by low temperature ( 9 ) . 0-8412-0473-X/80/33-188-211$06.75/l © 1980 American Chemical Society

Blank; Bioelectrochemistry: Ions, Surfaces, Membranes Advances in Chemistry; American Chemical Society: Washington, DC, 1980.

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4 msec

I

I 10.4

mV

msec Vision Research

Figure 1. The ERP of a cynomolgus monkey. Responses were recorded by a tungsten microelectroae at about the level of the inner segments in the retina, with the reference electrode in the vitreous humor of the eye. The stimulus was an intense 20-fisec flash. In Record A, the ERP intervenes between the stimulusflashand the rising phase of the a wave, which is part of the LRP. The time course of decay of the ERP, when isolated by cessation of artificial respiration, is shown by the dashed line. In Record B, the stimulus artifact consists only of the positive tip which is labeled. Thus the ERP is biphasic, and is followed by the LRP (32). The advent of techniques of forming model membrane systems makes it possible to study membrane phenomena under well-defined experimental conditions (10,11,12,13,14). Since Tien (15,16) discovered a photoelectric effect in a chlorophyll-containing (artificial) lipid bilayer (synonymous w i t h bilayer or bimolecular lipid membrane ( B L M ) ) , similar effects have been found and studied in lipid bilayers which contain other membrane-bound pigments. A fast photocurrent that possesses all the major characteristics of the E R P has been recorded i n a lipid bilayer which contains a magnesium porphyrin and separates a redox gradient (17,18). More recently, Montal and his co-workers succeeded in demonstrating an E R P - l i k e signal i n a rhodopsin-containing model membrane (19,20), as well as in a model membrane that contains bacteriorhodopsin from Halobacterium halobium (21). In this survey, we shall review selected data concerned with displacement photoelectric currents. Reviews on the photoelectric effects i n biomembranes (18,22-29) and on the E R P (5,30-37) are available. The data w i l l be analyzed i n terms of an equivalent circuit scheme, which


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Pigment-Containing Biomembranes

is generalized from model-system studies. W e shall distinguish primary photoelectric events from the secondary ionic events. W e further divide the primary effect into an ac (capacitative) photoelectric effect and a dc (resistive) photoelectric effect (28). Thus, the various descriptive terms such as fast photoelectric effect (fast photovoltage; F P V ) (38), photoelectric potentials ( P E P ) (39), and displacement photocurrents (40) w i l l be regarded as synonymous w i t h the ac photoelectric effect. W e shall describe two possible molecular models for the ac photoelectric effect (18,28,37), and discuss their validity i n visual and i n purple membranes i n terms of available experimental data. Finally, w e shall speculate on possible physiological significance of the ac photoelectric effect. Classification of Photoelectric Effects Operationally, a photoemf (photoelectromotive force) can be defined if stimulation by light generates an electric current across a pigment-containing membrane with two sides of the membrane kept equipotential. The term primary photoelectric effect refers to this current and the electrochemical gradient so generated. This definition thus excludes the secondary electrical events owing to dissipation of this gradient or a pre-existing gradient through light-controlled modulation of membrane ionic permeabilities. The latter are of considerable biological importance, but are beyond the scope of the present review (for a brief discussion and further references see Ref. 28). The primary photoelectric effect can be divided further into a dc and an ac effect. A dc photoresponse leads to a net transfer of charges across the membrane, while an ac photoresponse results i n no net transfer of charges. This classification is visualized best i n terms of a generalized equivalent circuit, which was proposed initially to account for quantitative data i n a model system of pigment-containing lipid bilayer (see Figure 2 ) . The experimental basis of this scheme has been published elsewhere (17,18, 28). In this generalized scheme, a parallel combination of ionic resistance, R , and membrane capacitance, C , represents a plain unmodified lipid bilayer. In addition, pigment photoreactions create a parallel electric pathway that includes a photoemf, E (with an internal resistance, fl ), which drives photoelectric current through another RC network: a transmembrane resistance, R , and a capacitance, C . The mechanism of charge transport across the membrane (through R ) can be quite different from the ionic mechanisms encountered in nerve impulse generation. This dc current generally is not carried by the inorganic ions ( N a , K , C l , etc.) that are present i n the aqueous phases i n relative abundance (18,41). Instead, the current may be carried either by mobile m

m

p

p

g

p

6

+

+

+


214


IM

Pigmented Membrane

Figure 2. A generalized equivalent circuit for a pigmented membrane and a measuring device. The photoreactions are represented by a photoemf, E^t), with an internal resistance R . The photoemf drives a dc photoelectric current through a transmembrane pigment resistance, R„ and an ac photoelectric current through a chemical capacitance, C . This photoelectric network is connected in parallel with the membrane ionic resistance, R , and the membrane capacitance, Cm. The measuring device has an access impedance, which is usually frequence dependent, but can be replaced by an effective access resistance, R„ which is constant within the bandwidth of interest. The access impedance is the sum of the input impedance of the measuring device, the electrode impedance, and the resistance owing to intervening electrolyte solutions. Thus the relationship holds: V(t) = I(t) R«. In an open-circuit measurement, l(t) is so small that V(t) is being read instead. The time course of l(t) is dependent on the value of R, (17J. p

9

m

charged pigment molecules (41) or by a solid-state mechanism involving an electron-hole production (22) or a proton hopping i n the pigment molecule (42,43). A t the two membrane-water interfaces, the continuity of a dc electric current requires that this specific charge-carrying function be relayed to the major inorganic ions i n the aqueous phases through an interfacial electron (or proton) transfer reaction. This interfacial charge transfer is usually not strictly one way and a reverse interfacial transfer can occur as the system relaxes. H o n g and Mauzerall (17,18) have shown that such an interfacial charge transfer and the subsequent reverse transfer are tantamount to charging and discharging a capacitance that is connected in series to the photoemf. This capacitance is named chemical capacitance to distinguish it from the membrane capacitance that is connected in parallel. Historically, this interfacial charge transfer process has never been given serious consideration as a possible generating mechanism of the ac photoelectric effect. Instead, the E R P , for example, is regarded generally as a manifestation of a light-induced internal charge separation (displacement) of the visual pigment rhodopsin. These two molecular mechanisms w i l l be described briefly later. For a detailed evaluation of these two mechanisms, interested readers are referred to Ref. 37.


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In a simple system such as the magnesium porphyrin-aqueous redox system studied by H o n g and Mauzerall (17,18), the resistive elements in the equivalent circuit can be treated as idealized elements with constant values, and all the RC parameters can be determined experimentally. The measured photocurrent can be fit to the computed time course w i t h the amplitude of the photoemf as the only freely adjustable parameter (Figure 3 A ) . Generally, the resistive elements need not be idealized elements. They can be voltage a n d / o r time dependent. The secondary ionic events are incorporated into a single resistance, R . W i t h this generalized model, we now can attempt to identify, at least tentatively, various signals in representative photobiological systems. Photosynthetic membranes in plants contain chlorophyll-protein complexes as reaction centers, and electron transport chain components (44,45). In higher plants, there are two reaction centers designated as photosystems I and II. Evidence indicates that these components are oriented and asymmetrically distributed across the photosynthetic membranes. Photons absorbed by the reaction centers initiate electron transfer reactions that are relayed through the electron transport chain. Eventually a proton gradient is generated across the photosynthetic membranes m

250 < 200

Z

150

• 100 o

o *» o

50 0

-50

50

100 Time

150

200

(us) Proceedings of the National Academy of Sciences of the USA

Figure 3. Record A. Photocurrent responses from a lipid bilayer that contains lipid-soluble magnesium meso-porphyrin di-n-amyl ester and separates a redox gradient with lOmM potassium ferricyanide and 0.5mM potassium ferrocyanide in one side (reference side) and lOmM potassium ferrocyaniae in the other side. The aqueous phase also contains I M NaCl buffered at pH 7.2. Temperature is 26°C. The light source is a dye laser pulse of 300-nsec duration and 590-nm output. The current was measured at an access impedance of 5.1 k& and an instrumental time constant of 1.5 usee: ( ), computed response based on the equivalent circuit shown in Figure 2, with input parameters determined experimentally; (- • •), the measured response which was an average of 16 measurements. Notice that the area bound by the positive peak equals that bound by the negative peak (zero time integral). The dc photocurrent is estimated to be less than 1 nA and was not detected in this measurement (17). Blank; Bioelectrochemistry: Ions, Surfaces, Membranes Advances in Chemistry; American Chemical Society: Washington, DC, 1980.

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IONS, SURFACES, M E M B R A N E S

Biochimica et Biophysica Acta

Figure 3. Record B. Open-circuit photovoltages across a lipid bilayer with a cyanine dye adsorbed to one interface. The light source is a xenon flash (8-fxsec duration) delivered at time indicated by the arrows. The left record is the control with an access impedance of lO'tt. The right record shows the effect of adding a shunt resistor of 10 Sl (58). 7

with the inside being more acidic than the outside of the photosynthetic membrane. Thus, photoreactions i n the two photosystems can be represented by two photoemfs connected in series that drive a dc photocurrent through the membrane. The dissipation of this gradient is coupled to the synthesis of A T P (46) i n accordance with Mitchell's chemiosmotic hypothesis (47) (secondary ionic event). As we shall see later i n this chapter, an ac photoelectric effect also might be present. This ac photoelectric effect can be identified with the fast light-induced voltages i n chloroplasts (48,49,50).

f-H- -hH-h - H - H - -H-H- -H-H '-H-H- +H+-H-H-4-H4-4+4 1


Figure 3. Record C. Photoelectric responses from a lipid bilayer containing retinal with 2 m M potassium ferricyanide added to one aqueous phase. Scale: 1 mV/division; 5 pA/division; 2 sec/division. The light pulse (top trace) has a duration of 8 sec and a rise time of 5 msec. The photovokage (middle trace) was measured by an electrometer (input impedance, 10 $l; rise time, 200 msec). The photocurrent (bottom trace) was measured by a picoammeter (input impedance, I0 &; rise time, 20 msec) (57). 16

7


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*

217

•

f f 1 f


Figure 3. Record D. Photoelectric responses from a livid bilayer containing Vitamin A acid with 2mM potassium ferricyaniae added to one aqueous phase. Scale: 0.2 mV/division; 10 pAIdivision; 2 sec/division. The measurements of the light pulse (top trace), the photovoltage (middle trace), and the photocurrent (bottom trace) are similar to those in Record C (57). In the visual photoreceptor membranes, no dc photoelectric signal has ever been demonstrated convincingly. The two components R l and R2 of the E R P are ac photoelectric signals (4,51) and the photoemfs for these two components may be regarded as two consecutive photoemfs of opposite polarities (37). These two photoemfs drive electric current through a chemical capacitance, C , but not through a resistance, R (effectively, R = infinity). Montal et al. (52) have presented evidence of light-induced ionic channel formation i n reconstituted rhodopsin lipid bilayer membranes. The electric current i n this light-modulated channel must be regarded then as a secondary ionic event, according to the present classification, i.e. the current flow through R . A photobiological system which has received much attention i n the past few years is the purple membrane of Holobacterium halobium (53). The purple membrane contains only one single species of protein, bacteriorhodopsin, which contains retinal as the chromophore, similar to the visual pigment rhodopsin. However, it functions as a fight-driven proton pump (54). The resulting proton gradient is utilized i n the synthesis of A T P (55) similar to a photosynthetic membrane. The lightinduced proton current across the purple membrane is a dc photoelectric current, while the passive dissipation of the proton gradient must be regarded as a secondary ionic event. The presence of an ac photoelectric effect was suggested first by Hong (28) and demonstrated experimentally by Trissl and Montal (21). p

8

s

m


218



UmV

9

-3

2 min

-6

FEBS Letters

Figure 3. Record E. Open-circuit photovoltages across a lipid membrane which contains oriented bacteriorhodopsin. The two aqueous phases have identical compositions: 0.15M KCl and lOmM Tris-HCl at pH 6.3: (top trace), the control photovoltage; (bottom trace) the photovoltage after adding 2 X 10" M of CCCP. Shunting with an external resistor has a similar effect (59). 7

Methods of Measuring ac Photoelectric Responses A systematic discussion of the methods of measuring the photoresponses from a typical photosystem as represented by the equivalent circuit i n Figure 2 w i l l facilitate our interpretation and classification of various photoelectric data. L e t us consider the two common methods of electric measurements: galvanostatic (also known to electrophysiologists as the current clamp or open-circuit method) and potentiostatic (also known as the voltage clamp or short-circuit method). I n an idealized galvanostatic measurement, the measuring device should have an infinite input impedance (relative to the source impedance of the system being


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measured) so that no appreciable current is drawn by the measuring device. In an idealized potentiostatic measurement, the two aqueous phases near each membrane-water interface should be maintained at a fixed potential at all time, and thus the access impedance, Re, (sum of the input impedance of the amplifier, the electrode impedance, and the impedance caused by intervening electrolyte solutions) must be effectively zero relative to the membrane source impedance. It is instructive to consider the effect of varying the access impedance and the effect of a shunting path such as produced by adding ionophores to the membrane or produced by connecting a parallel resistor across the input of the amplifier.

Control

r

t 10

VMC.

30

mM

K

t

ft Plant Science Letters

Figure 3. Record F. Open-circuit photovoltages recorded intracellularly from an intact chloroplast of Peperomia metallica. Open and closed arrows mark the "on" and the "off,' respectively, of the rectangular pulses of white light Left record is the control in the absence of valinomycin. Right record shows the effect of 1/zM valinomycin and 30mM external potassium ions. The shunting effect of valinomycin shortens the relaxation time constants and reduces the amplitude of the steady-state photovoltage in accordance with the equivalent circuit shown in Figure 2 (67).


220


In a steady-state galvanostatic measurement, the effect of shunting is a reduction in the amplitude of the photoelectric signal. Thus, a very "leaky" membrane presents a difficult condition for measuring a photovoltage. In contrast, the steady-state dc photoelectric current, as measured by a (quasi-) potentiostatic method, remains essentially constant when the ionic resistance (R ) is decreased by several orders of magnitude (18,41). In fact, this method can be used to differentiate between the primary photoelectric and the secondary ionic events. The effect of shunting on transient photoelectric signals is more complex, since both the amplitude and the relaxation time course of the photosignals may be affected. W e shall consider a type of measurement named the tunable voltage clamp method, first introduced by Hong and Mauzerall (17, 56), in which a finite and nonzero access impedance is included in the data analysis. The galvanostatic and potentiostatic measurements can be regarded then as two extreme special cases with the access impedance approaching infinity and zero, respectively. In a photosystem i n which all of the RC elements are constant (voltage independent and time i n variant), it is possible to obtain analytical results explicitly. Here we shall summarize the results with respect to two types of light stimulation, namely, brief pulse stimulation, ( i n which the duration of the light pulse is much shorter than the intrinsic relaxation time of the membrane photosystem), and the long rectangular pulse stimulation (in which the duration of the fight pulse is much longer than the intrinsic relaxation time of the photosystem) (see Figure 4 ) . The analytical details can be found in Ref. 18. Two important parameters deserve attention: a membrane discharging time constant ( r ) and an intrinsic photoelectric relaxation time constant ( T ) ; m

m

P

l

-1--I- + -J -

(i)

and ± - - ± -

+ - L -

(2)

Except for the case of an idealized potentiostatic measurement, the measured responses are always the results of interactions between T and T and other RC time constants, resulting in two apparent relaxation time constants, T (the short apparent relaxation time constant) and n (the long apparent relaxation time constant), which are dependent on the value of the access impedance in a way that is predictable by the equivalent circuit. Such a relationship has been confirmed quantitatively in magnesium porphyrin-containing lipid bilayers (see Figure 3 A ) . In the limit of infinite access impedance (galvanostatic), n approaches the p

M

8



m c

e

R »0

m

>r >o

oo >

r

e

R> 0

m

r =o

e

R = 0

m

M C

EXISTS

Ep(t)

m

_nr-L

LONG RECTANGULAR PULSE STIMULATION

SPECIAL CASE IN WHICH NO

Ep(t) = S(t).Ep

BRIEF PULSE STIMULATION

p

Figure 4. The dependence of time course of photoelectric responses on the membrane discharging time constant. Two types of stimulations are considered: brief pulse stimulation and long rectangular pulse stimulation: T is the intrinsic relaxation time constant: r is the membrane discharging time constant; r, and n are the short and long apparent relaxation time constants, respectively; and T is a critical value of r . See text for further explanation.

—•

*5

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membrane time constant, R C , while T is aproximately twice the value of T (18). ( O f course, i n a galvanostatic measurement, the current through the measuring device is effectively zero and the voltage, which is proportional to the small residual current, is measured instead.) Frequently r is masked completely because of inadequate time resolution (T is usually i n the microsecond range). In this case, the galvanostatic measurement could be devoid of relevant kinetic information, since it would measure nothing but the membrane R C relaxation. Failure to recognize this fact can lead to an erroneous interpretation of experimental data. In view of this distorting effect of a nonzero access impedance, one might prefer to consider an ideal potentiostatic measurement instead. However, a zero access impedance is very difficult, if not impossible, to achieve. H o n g and Mauzerall (18, 56) have shown that an access impedance of 380 O is still far from effectively zero at 1 M H z , mainly because of the presence of C , which bestows the membrane system with a low source impedance (of the order of 10 kft at 1 M H z ) . Furthermore, a nonzero but small access impedance can be a blessing rather than a nuisance, since additional information that is inaccessible i n an ideal potentiostatic measurement can be obtained. It has been suggested that the optimal measurement is that w i t h the access impedance tuned to match the source impedance (hence the name tunable voltage clamp method). m

m

b

p

8

P

p

Caution must be exercised i n using an instrument to measure a (quasi-) short-circuit current. A crucial parameter is the input impedance of the instrument i n addition to the electrode impedance. Since the source impedance of the membrane can be as low as 10 ko at 1 M H z , it is not good enough to have an input impedance of 10 O (that of a commercial picoammeter). It has been reported that the photocurrent response is the first time derivative of the photovoltage response (19,20). However, this type of relationship possibly indicates that the measurement was not made under short-ciicuit conditions. It is true that current flow through a discharging capacitor is proportional to the first time derivative of the voltage across it. It is equally true that the discharging of a capacitor is accelerated when it is connected to a low-input i m pedance device ( i n effect, reducing the discharging time constant). Therefore, the first time derivative relationship holds only when both the current and the voltage are measured under conditions with the same access impedance (relative to the source impedance) and should break down otherwise. Without this precaution in mind, a current measurement could be misleading. 7

The diagram i n Figure 4 indicates that as r is changing from an idealized zero to its maximum value R C , the photoelectric relaxation time course also changes continuously, w i t h increasing distortion owing m

m

m


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to interaction with the other RC parameters, which are unrelated to the photoelectric event. In general, a critical value ( T ) of the discharging time constant exists. If r < T , the brief pulse response is characterized by a biphasic waveform (a positive spike followed by an undershoot below the dark baseline), while the long rectangular pulse response is characterized by an "on-overshoot" and an "off-undershoot." I n addition, a dc component also may be present. The net amount of charges transferred across the membrane is represented by the time integral of the current response. In a measurement with a short r , the ac component is much enhanced and the time integral of the current response becomes almost zero (the amplitude of the dc component is negligible) (see Figure 3 A ) . As r becomes longer, the ac component decreases i n amplitude progressively, owing to increasing distortion through interaction of various RC constants. Eventually, when r exceeds the critical value T , the brief pulse response relaxes monotonically to the baseline without an undershoot while the long pulse response rises and decays monotonically. W e shall refer to the "undershoot" i n brief pulse responses and the "on-overshoot" and "off-undershoot" i n long pulse responses as the characteristic high-pass-filter response, since this waveform is characteristic of a high-pass RC filter. Thus, when r is reduced by the shunting effect of ionophores or a parallel shunt resistor, or by a decrease i n access impedance, a characteristic high-pass-filter response might appear. In Figure 3C, using a picoammeter (input impedance 10 ft) i n place of an electrometer (input impedance 10 O) causes a sufficient reduction of the access impedance (and hence the discharging time constant) to reveal the characteristic high-pass-filter response i n a lipid bilayer which contains retinal (57). Other examples are given by a cyanine-dye-containing l i p i d bilayer (58), which is shunted by a resistor of 10 Q (see Figure 3 B ) , and by a bacteriorhodopsin-containing model membrane (59,60, 61), which is shunted by an ionophore C C C P (2,4,6-trichlorocarbonylcyanidephenylhydrazone) (see Figure 3 E ) . Results similar to Figure 3 E are found also i n model membranes, which contain subchromatophore pigment-protein complexes from Rhodospirillum rubrum (62,63,64). Usually the appearance of a characteristic high-pass-filter response can be construed as the presence of a series capacitance connected to the photoemf, namely, a chemical capacitance. However the data presented in Refs. 62, 63, and 64 require a special consideration. T h e membranes were made by fusions of pigment-containing lipid vesicles to a plain planar membrane. The effect of ionophores and a shunt resistor has been interpreted as a consequence of a peculiar structure i n which bacteriorhodopsin-lipid vesicles remain intact but attached to a planar membrane (see Figure 12 i n Ref. 63). Thus there are three membrane capacitances: one of the planar membrane, one of the vesicle, and one of the M C

m

MC

m

m

m

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m

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fused region. The interaction among these three capacitances qualitatively explains the observed effect. In other words, the capacitance at the fused region fulfills the requirement of a series capacitance. H o w ever, this interpretation has an inherent difficulty. Here, it must be pointed out that the characteristic high-pass-filter response appears also in membranes made by a conventional method i n which no fusion with vesicles took place (59,65). The simplest interpretation is that the characteristic high-pass-filter response is a manifestation of an ac photoelectric effect (28) rather than an artifact specific to the particular method of making pigment-containing membranes. There are, however, special cases i n which a critical discharging time constant ( T ) does not exist (e.g., when R = infinity). In these cases, the characteristic high-pass-filter response persists even under opencircuit conditions (see e.g. Figures 3 D and 3F cited here and Figure 4 in Ref. 66). Here, the data obtained from the chloroplast of Peperomia metallica (67) shown in Figure 3F suggest the existence of an ac photoelectric signal. M C

a

It is apparent that the equivalent circuit shown i n Figure 2 offers a simple and unified interpretation of ac photoresponses from different systems. F o r example, the difference of photoresponses shown i n Figures 3C and 3 D merely reflects the difference i n relative magnitude of various RC parameters in the equivalent circuit (18); there is no need to postulate different models to explain the results (57). Molecular Basis of a Chemical Capacitance W e have emphasized that the ac photoelectric response is a manifestation of a chemical capacitance, which is physically distinct from the membrane capacitance; the former is connected in series to the photoemf while the latter is connected in parallel. W e shall briefly present molecular pictures to demonstrate that this is indeed the case. W e shall summarize only the basic feature of two molecular models which are results of a calculation based on the Gouy-Chapman double-layer theory. The interested readers are referred to Refs. 18 and 37 for the details. Interfacial Charge Transfer (ICT) Model. The basic feature of this model is that the membrane-bound pigment can reversibly exchange a charge (electron or proton) with the aqueous charge donors or acceptors upon light illumination. Since there are two interfaces and electric signals generated there tend to cancel each other, no external detection can be made unless an asymmetry exists. This asymmetry may be in the two aqueous phases (e.g., a redox or p H gradient) or i n the membrane (e.g., an asymmetric and oriented distribution of photopigment i n the membrane). W e shall consider a simple case in which interfacial charge-


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transfer reactions occur at a single membrane-water interface (see the left interface i n Figure 5 ) . The interfacial charge-transfer reactions are usually faster than the transmembrane charge transport (dc effect) and thus the latter can be ignored during the brief transient of an ac photoresponse. The light-induced interfacial charge transfer and its subsequent relaxation thus causes formation and decay of a sheet of surface charge at the interface. A simple calculation (18) shows that the ionic atmosphere relaxation time is less than 1 nsec i n an isotonic electrolyte solution, while it can be as long as 1 sec i n the membrane phase. W e assume that the interfacial charge recombination occurs on a time scale of 1 fisec to 1 msec. Thus, the ionic distribution i n the aqueous phase is always i n quasiequilibrium during the transient, while there is practically no charge screening i n the membrane phase. This feature makes it possible to carry out the kinetic and the electrostatic calculations independently and separately. Also a constant field condition can be imposed i n the membrane phase because of insignificant space charge build-up i n the membrane phase. As a result of such a calculation, the potential profile and the charge-density profile across the membrane and the aqueous phases under short-circuit conditions are shown in Figure 5 (see the left half of the figure), which must be regarded as a snapshot during the brief transient of the ac photoresponse. As expected the charged pigment molecules are concentrated near the reacting interface. Because of the extremely small thickness ( < 7 0 A ) of a membrane, the polarizing effect of the surface charge extends beyond the membrane phase and reaches the opposite aqueous phase. Thus, the positive surface charge is balanced by the excess negative charges at both diffuse double layers under short-circuit conditions. Linearization of the results permits a re-interpretation of the profiles i n terms of three lumped capacitances: a geometric capacitance, C , owing to the membrane dielectric and two double-layer capacitances, C . Notice that the double-layer capacitance at the reacting interface has an opposite polarity to the remaining two capacitances. The only sensible way to connect a photoemf i n order to generate such a charge distribution is as shown i n Figure 5 (see the left half of the figure). Macroscopically, this ac photoresponse is characterized by a high-pass-filter circuit with a series capacitance, C . Here C is the result of a combination of three more fundamental capacitances: the geometric capacitance and the opposite double-layer capacitance combine i n series; the result of such a combination then combines i n parallel with the adjacent double-layer capacitance. C is thus physically distinct from the membrane capacitance, which is a series combination of the same three fundamental capacitances. g

d

p

p

p

Oriented Dipole Model. The basic feature of this model is that the membrane-bound pigment is oriented and distributed asymmetrically across the membrane, and that a light-induced conformational change


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ICT MODEL

OD MODEL

P

hv

Y

aqueous membrane aqueous

4—

Displacement Photocurrents in Pigment-Containing Biomembranes

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