ATP-powered IC

Figure 1a shows the complete hybrid integrated system, consisting of a CMOS IC packaged with an ATP-harvesting ‘biocell’. The biocell consists of two series-stacked ATPase bearing suspended lipid bilayers with a fluid chamber directly on top of the IC. Series stacking of two membranes is necessary to provide the required start-up voltage for IC and eliminates the need for an external energy source, which is typically required to start circuits from low-voltage supplies2,3. As shown in Fig. 1c, a matching network in the form of a switched capacitor allows the load resistance of the IC to be matched to that presented by the biocell. In principle, the switch S can be implicit. The biocell charges C STOR until the self start-up voltage, V start , is reached. The chip then operates until the biocell voltage drops below the minimum supply voltage for operation, V min . Active current draw from the IC stops at this point, allowing the charge to build up again on C STOR . In our case, however, the IC leakage current exceeds 13.5 nA at V start , more than can be provided by the biocell. As a result, an explicit transistor switch and comparator (outside of the IC) are used for this function in the experimental results presented here, which are not powered by the biocell and not included in energy efficiency calculations (see Supplementary Discussion for additional details). The energy from the biocell is used to operate a voltage converter (voltage doubler) and some simple inverter-based ring oscillators in the IC, which receive power from no other sources.

Figure 1: Fully hybrid biological–solid-state system. (a) Illustration depicting biocell attached to CMOS integrated circuit. (b) Illustration of membrane in pore containing sodium–potassium pumps. (c) Circuit model of equivalent stacked membranes, =2.1 pA, =98.6 GΩ, =575 GΩ and =75 pF, Ag/AgCl electrode equivalent resistance R WE +R CE <20 kΩ, energy-harvesting capacitor C STOR =100 nF combined with switch as an impedance transformation network (only one switch necessary due to small duty cycle), and CMOS IC voltage doubler and resistor representing digital switching load. R L represents the four independent ring oscillator loads. (d) Equivalent circuit detail of stacked biocell. (e) Switched-capacitor voltage doubler circuit schematic. Full size image

A silver/silver-chloride (Ag/AgCl) microfabricated thin film on the surface of the chip, and an Ag/AgCl pellet serve as the working (WE) and counter (CE) electrode to convert ions to electrons. Careful attention must be paid to the electrodes in this power-transfer application to ensure that the electrodes are not an uncontrolled galvanic energy source. The lipid (purified 1,2-dioleoyl-sn-glycero-3-phosphocholine, DOPC) bilayers are formed in a 250-μm pore12 epoxied to a teflon housing (see Methods for details). Purified 5′-sodium potassium adenosine triphosphatase from porcine cerebral cortex is embedded into each bilayer (see Methods for details). Prior to the addition of ATP, the membrane produces no electrical power and has an R m of 280 GΩ. A 1.7-pA short-circuit (SC) current (Fig. 2b) through the membrane is observed upon the addition of ATP (final concentration 3 mM) to the cis chamber where functional, properly oriented enzymes generate a net electrogenic pump current. To perform these measurements, currents through each membrane of the biocell are measured using a voltage-clamp amplifier (inset of Fig. 2b) with a gain of 500 GΩ with special efforts taken to compensate amplifier leakage currents. Each ATPase transports three Na+ ions from the cis chamber to the trans chamber and two K+ ions from the trans chamber to the cis chamber (a net charge movement of one cation) for every molecule of ATP hydrolysed. At a rate of 100 hydrolysis events per second under zero electrical (SC) bias13, this results in an electrogenic current of ∼16 aA. The observed SC current corresponds to about 105 active ATPases in the membrane or a concentration of about 2 × 106 mm−2, about 5% of the density of channels occurring naturally in mammalian nerve fibres14. It is expected that half of the channels inserted are inactive because they are oriented incorrectly.

Figure 2: Single-cell biocell characterization. (a) I–V sweeps of biocell before (▴) and after (●) addition of ATP (3 mM final concentration). Voltages are stepped for this measurement. Bilayer current is the average of the last 5 s of a 10-s step (after charging currents have decayed away). Bilayer capacitance C m =153 pF (consistent with DOPC bilayers12,30). Pre-ATP data linear fit (black line) slope yield R m =280 GΩ. Post ATP data fit to a Boltzmann curve, slope=0.02 V (blue line). Post-ATP linear fit (red line) yields I p =−1.8 pA and R p =61.6 GΩ, which corresponds to a per-ATP source resistance of 6.16 × 1015. The current due to membrane leakage through R_{m} is subtracted in the post-ATP curve. (b) Short circuit current magnitude increasing to 1.7 pA after the addition of 3 mM ATP. Transimpedance amplifier gain is 500 GΩ. The headstage filter consists of a three-pole analog low-pass Bessel filter f c =10 kHz in series with four-pole analog low-pass Bessel filter f c =100 Hz. Data is post-processed with a low-pass digital filter with f c =1 Hz. (c) Biocell charging its own capacitance to 78 mV. Measured (black line). Simulated using non-linear model fit (blue line). Full size image

Current–voltage characteristics of the ATPases

Figure 2a shows the complete measured current–voltage (I–V) characteristic of a single ATPase-bearing membrane in the presence of ATP. The current due to membrane leakage through R m is subtracted in the post-ATP curve. The I–V characteristic fits a Boltzmann sigmoid curve, consistent with sodium–potassium pump currents measured on membrane patches at similar buffer conditions13,15,16. This nonlinear behaviour reflects the fact that the full ATPase transport cycle (three Na+ ions from cis to trans and two K+ ions from trans to cis) time increases (the turn-over rate, k ATP , decreases) as the membrane potential increases16. No effect on pump current is expected from any ion concentration gradients produced by the action of the ATPases (see Supplementary Discussion). Using this Boltzmann fit, we can model the biocell as a nonlinear voltage-controlled current source I ATPase (inset Fig. 2a), in which the current produced by this source varies as a function of V m . In the fourth quadrant, where the cell is producing electrical power, this model can be linearized as a Norton equivalent circuit, consisting of a DC current source (I p ) in parallel with a current-limiting resistor (R p ), which acts to limit the current delivered to the load at increasing bias (I ATPase ∼I p −V m /R p ). Figure 2c shows the measured and simulated charging of C m for a single membrane (open-circuited voltage). A custom amplifier with input resistance R in >10 TΩ was required for this measurement (see Electrical Measurement Methods).

Reconciling operating voltage differences

The electrical characteristics of biological systems and solid-state systems are mismatched in their operating voltages. The minimum operating voltage of solid-state systems is determined by the need for transistors to modulate a Maxwell–Boltzmann (MB) distribution of carriers by several orders of magnitude through the application of a potential that is several multiples of kT/q (where k is Boltzmann’s constant, T is the temperature in degrees Kelvin and q is the elementary charge). Biological systems, while operating under the same MB statistics, have no such constraints for operating ion channels since they are controlled by mechanical (or other conformational) processes rather than through modulation of a potential barrier. To bridge this operating voltage mismatch, the circuit includes a switched-capacitor voltage doubler (Fig. 1d) that is capable of self-startup from voltages as low V start =145 mV (∼5.5 kT/q) and can be operated continuously from input voltages from as low as V min =110 mV (see Supplementary Discussion). In the series-stacked biocell with each membrane having approximately equal resistance and capacitance, and . Once the converter has started up, operation can be maintained down to 110 mV, which means that each membrane in the two-stacked configuration can be biased to operate at potentials as low as 55 mV.

Maximizing the efficiency of harvesting energy from ATP

Solid-state systems and biological systems are also mismatched in their operating impedances. In our case, the biocell presents a source impedance, =84.2 GΩ, while the load impedance presented by the complete integrated circuit (including both the voltage converter and ring oscillator loads) is approximately R IC =200 kΩ. (The load impedance, R L , of the ring oscillators alone is 305 kΩ.) This mismatch in source and load impedance is manifest in large differences in power densities. In general, integrated circuits, even when operated at the point of minimum energy in subthreshold, consume on the order of 10−2 W mm−2 (or assuming a typical silicon chip thickness of 250 μm, 4 × 10−2 W mm−3) (ref. 17). Typical cells, in contrast, consume on the order of 4 × 10−6 W mm−3 (ref. 18). In our case, a typical active power dissipation for our circuit is 92.3 nW, and the active average harvesting power is 71.4 fW for the biocell. This discrepancy is managed through duty-cycled operation of the IC in which the circuit is largely disabled for long periods of time (T charge ), integrating up the power onto a storage capacitor (C STOR ), which is then expended in a very brief period of activity (T run ), as shown in Fig. 3a.

Figure 3: Complete hybrid biological–solid-state system characterization. (a) Duty-cycled operation of the stacked biocell and CMOS IC. Shown are the input (V m ) and output (V out ) voltage of the voltage doubler. T charge =6250, s, T run =5.5 ms. During T run , 54.4 nW delivered to the load and 37.9 nW is consumed in running the voltage doubler. (b) Expanded waveform showing V m and V out near a switching transition and one of four ring oscillators. Inset shows the output of ring oscillator digital switching load switching at 1 MHz. (c) Biocell current–voltage characteristic and effective load line. Full size image

The overall efficiency of the system in converting chemical energy to the energy consumed in the load ring oscillator (η) is given by the product of the conversion efficiency of the voltage doubler (η converter ) and the conversion efficiency of chemical energy to electrical energy in the biocell (η biocell ), η=η converter × η biocell . η converter is relatively constant over the range of input voltages at ∼59%, as determined by various loading test circuits included in the chip design (Supplementary Figs 1–6). η biocell , however, varies with transmembrane potential V m . η is the efficiency in transferring power to the power ring oscillator loads from the ATP harvested by biocell.

As reflected in the single-membrane circuit model of Fig. 4a, V m in cells is the result of two related potentials, both derived from the action of the ion pumps: the potential that develops as a result of ionic concentration gradients of membrane permeable ions (the Nernst, or diffusion, potential, V diff ) and Ohmic potentials (V p ) due to the net electrogenic ionic currents of the ion pumps. These combine to determine the membrane potential, V m =V diff +V p . V diff is determined by the superposition of the Thevenin ‘Nernst’ sources for each ionic species, , , , , where , R is the gas constant, z is the valence of the species, F is Faraday’s constant and [X] y is the concentration of species X in chamber y. A corresponding series resistance R X models the membrane permeability of those ions present in the system. An equivalent expression for V diff is also given by the Goldman–Hodgkin–Katz (GHK) equation. The total membrane resistance R m is given by the parallel combination of , , , . Using the linear ATPase model (Fig. 1c), the electrogenic component of the transmembrane potential is given by V p =I p (R p ||R m ). Under physiological conditions, V p is often negligible compared with V diff as a result of the relatively low value of R m due to the presence of many transmembrane ion channels.

Figure 4: Equivalent circuit models. (a) Single-biocell equivalent circuit model with additional voltage sources to account for energy delivered by the pump. (b) Stacked membrane equivalent circuit model. Full size image

To first order, the energy made available to the Na+/K+-ATPase by the hydrolysis of ATP is independent of the chemical or electric potential of the membrane and is given by |ΔG ATP |/(qN A ), where ΔG ATP is the Gibbs free energy change due to the ATP hydrolysis reaction per mole of ATP at given buffer conditions and N A is Avogadro’s number. Since every charge that passes through I ATPase corresponds to a single hydrolysis event, we can use two voltage sources in series with I ATPase to independently account for the energy expended by the pumps both in moving charge across the electric potential difference and in moving ions across the chemical potential difference. The dependent voltage source V loss in this branch fixes the voltage across I ATPase , and the total power produced by the pump current source is (|ΔG ATP |/N A )(Nk ATP ), which is the product of the energy released per molecule of ATP, the number of active ATPases and the ATP turnover rate. The power dissipated in voltage source V chem models the work performed by the ATPases in transporting ions against a concentration gradient. In the case of the Na+/K+ ATPase, V chem is given by . The power dissipated in this source is introduced back into the circuit in the power generated by the Nernst independent voltage sources, and . The power dissipated in the dependent voltage source V loss models any additional power not used to perform chemical or electrical work.

In our system, no significant ion concentration gradient exists, V chem =0, and the expanded membrane model of Fig. 4a becomes the passive membrane model of Fig. 4b. The efficiency of the biocell is given by with (the factor of two arises from the series stack, see Supplementary Discussion). The maximum efficiency occurs at . In our case, and the overall energy efficiency of the system η=η converter × η biocell =8.84%.

Higher membrane potentials can be sustained in systems with electrogenic ion pumps engineered with lower V chem . For example, proton-pumping ATPases, such as those found in the plasma membrane of the yeast Schizosaccharomyces pombe, transport only a single proton per hydrolysed molecule of ATP. Less chemical work is performed (V chem =14 mV, for ΔpH=0.1) compared with the sodium–potassium pumps. As a result, these pumps can sustain a membrane potential of up to 280 mV19,20. In our system, with a similar number of active incorporated pumps and R m in excess of 575 GΩ due to the absence of other channels, voltages of 227 mV could have been achieved with these pumps.