RESUMO
Recent years have seen the development of a number of biosensor architectures that rely on target binding-induced changes in the rate of electron transfer from an electrode-bound receptor. Most often, the interrogation of these sensors has relied on voltammetric methods, such as square-wave voltammetry, which limit their time resolution to a few seconds. Here, we describe the use of an impedance-based approach, which we have termed electrochemical phase interrogation, as a means of collecting high time resolution measurements with sensors in this class. Specifically, using changes in the electrochemical phase to monitor target binding in an electrochemical-aptamer based (EAB) sensor, we achieve subsecond temporal resolution and multihour stability in measurements performed directly in undiluted whole blood. Electrochemical phase interrogation also offers improved insights into EAB sensors' signaling mechanism. By modeling the interfacial resistance and capacitance using equivalent circuits, we find that the only parameter that is altered by target binding is the charge-transfer resistance. This confirms previous claims that binding-induced changes in electron-transfer kinetics drive signaling in this class of sensors. Considering that a wide range of electrochemical biosensor architectures rely on this signaling mechanism, we believe that electrochemical phase interrogation may prove generalizable toward subsecond measurements of molecular targets.
Assuntos
Aptâmeros de Nucleotídeos/química , Tobramicina/sangue , Técnicas Biossensoriais/instrumentação , Técnicas Biossensoriais/métodos , Técnicas Eletroquímicas , Hexanóis/química , Cinética , Modelos Químicos , Conformação Molecular , Oxirredução , Compostos de Sulfidrila/química , Propriedades de SuperfícieRESUMO
The development of a technology capable of tracking the levels of drugs, metabolites, and biomarkers in the body continuously and in real time would advance our understanding of health and our ability to detect and treat disease. It would, for example, enable therapies guided by high-resolution, patient-specific pharmacokinetics (including feedback-controlled drug delivery), opening new dimensions in personalized medicine. In response, we demonstrate here the ability of electrochemical aptamer-based (E-AB) sensors to support continuous, real-time, multihour measurements when emplaced directly in the circulatory systems of living animals. Specifically, we have used E-AB sensors to perform the multihour, real-time measurement of four drugs in the bloodstream of even awake, ambulatory rats, achieving precise molecular measurements at clinically relevant detection limits and high (3 s) temporal resolution, attributes suggesting that the approach could provide an important window into the study of physiology and pharmacokinetics.
Assuntos
Preparações Farmacêuticas/sangue , Preparações Farmacêuticas/metabolismo , Bibliotecas de Moléculas Pequenas/metabolismo , Animais , Aptâmeros de Nucleotídeos/metabolismo , Técnicas Biossensoriais/métodos , Bovinos , Humanos , Limite de Detecção , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Electrochemical sensors are major players in the race for improved molecular diagnostics due to their convenience, temporal resolution, manufacturing scalability, and their ability to support real-time measurements. This is evident in the ever-increasing number of health-related electrochemical sensing platforms, ranging from single-measurement point-of-care devices to wearable devices supporting immediate and continuous monitoring. In support of the need for such systems to rapidly process large data volumes, we describe here an open-source, easily customizable, multiplatform compatible program for the real-time control, processing, and visualization of electrochemical data. The software's architecture is modular and fully documented, allowing the easy customization of the code to support the processing of voltammetric (e.g., square-wave and cyclic) and chronoamperometric data. The program, which we have called Software for the Analysis and Continuous Monitoring of Electrochemical Systems (SACMES), also includes a graphical interface allowing the user to easily change analysis parameters (e.g., signal/noise processing, baseline correction) in real-time. To demonstrate the versatility of SACMES we use it here to analyze the real-time data output by (1) the electrochemical, aptamer-based measurement of a specific small-molecule target, (2) a monoclonal antibody-detecting DNA-scaffold sensor, and (3) the determination of the folding thermodynamics of an electrode-attached, redox-reporter-modified protein.
Assuntos
Eletroquímica/métodos , Software , Animais , Aptâmeros de Nucleotídeos/genética , Aptâmeros de Nucleotídeos/metabolismo , Sequência de Bases , Técnicas Biossensoriais , DNA/análise , Processamento Eletrônico de Dados , Masculino , Dobramento de Proteína , Ratos , Ratos Sprague-Dawley , Razão Sinal-Ruído , Fatores de TempoRESUMO
The electrochemical, aptamer-based (E-AB) sensor platform provides a modular approach to the continuous, real-time measurement of specific molecular targets (irrespective of their chemical reactivity) in situ in the living body. To achieve this, however, requires the fabrication of sensors small enough to insert into a vein, which, for the rat animal model we employ, entails devices less than 200 µm in diameter. The limited surface area of these small devices leads, in turn, to low faradaic currents and poor signal-to-noise ratios when deployed in the complex, fluctuating environments found in vivo. In response we have developed an electrochemical roughening approach that enhances the signaling of small electrochemical sensors by increasing the microscopic surface area of gold electrodes, allowing in this case more redox-reporter-modified aptamers to be packed onto the surface, thus producing significantly improved signal-to-noise ratios. Unlike previous approaches to achieving microscopically rough gold surfaces, our method employs chronoamperometric pulsing in a 5 min etching process easily compatible with batch manufacturing. Using these high surface area electrodes, we demonstrate the ability of E-AB sensors to measure complete drug pharmacokinetic profiles in live rats with precision of better than 10% in the determination of drug disposition parameters.
Assuntos
Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais , Técnicas Eletroquímicas , Animais , Eletrodos , Tamanho da Partícula , Ratos , Propriedades de SuperfícieRESUMO
Anomalies in prefrontal cortex (PFC) function are posited to underpin difficulties in learning to suppress drug-seeking behavior during abstinence. Because group 1 metabotropic glutamate receptors (mGluRs) regulate drug-related learning, we assayed the consequences of extended access to intravenous cocaine (6 h/d; 0.25 mg/infusion for 10 d) on the PFC expression of group 1 mGluRs and the relevance of observed changes for cocaine seeking. After protracted withdrawal, cocaine-experienced animals exhibited a time-dependent intensification of cue-induced cocaine-seeking behavior and an impaired extinction of this behavior. These behavioral phenomena were associated with a time-dependent reduction in mGluR1/5 expression within ventromedial PFC (vmPFC) of cocaine-experienced animals exposed to extinction testing but not in untested ones. Interestingly, pharmacological manipulations of vmPFC mGluR1/5 produced no immediate effects on cue-induced cocaine-seeking behavior but produced residual effects on a subsequent test for cocaine seeking. At 3 d withdrawal, cocaine-experienced rats infused intra-vmPFC with mGluR1/5 antagonists, either before or after an initial test for cocaine seeking, persisted in their cocaine seeking akin to cocaine-experienced rats in protracted withdrawal. Conversely, cocaine-experienced rats infused with an mGluR1/5 agonist before the initial test for cocaine-seeking at 30 d withdrawal exhibited a facilitation of extinction learning. These data indicate that cue-elicited deficits in vmPFC group 1 mGluR function mediate resistance to extinction during protracted withdrawal from a history of extensive cocaine self-administration and pose pharmacological stimulation of these receptors as a potential approach to facilitate learned suppression of drug-seeking behavior that may aid drug abstinence.
Assuntos
Transtornos Relacionados ao Uso de Cocaína/fisiopatologia , Extinção Psicológica/fisiologia , Córtex Pré-Frontal/fisiologia , Receptores de Glutamato Metabotrópico/fisiologia , Síndrome de Abstinência a Substâncias/fisiopatologia , Animais , Western Blotting , Transtornos Relacionados ao Uso de Cocaína/psicologia , Condicionamento Operante , Sinais (Psicologia) , Antagonistas de Aminoácidos Excitatórios/farmacologia , Extinção Psicológica/efeitos dos fármacos , Injeções , Masculino , Metoxi-Hidroxifenilglicol/análogos & derivados , Metoxi-Hidroxifenilglicol/farmacologia , Córtex Pré-Frontal/efeitos dos fármacos , Piridinas/administração & dosagem , Piridinas/farmacologia , Ratos , Ratos Sprague-Dawley , Receptores de Glutamato Metabotrópico/biossíntese , Receptores de Glutamato Metabotrópico/efeitos dos fármacos , Recidiva , Autoadministração , Síndrome de Abstinência a Substâncias/psicologiaRESUMO
Histone deacetylase inhibitors (HDACIs) strengthen memory following fear conditioning and cocaine-induced conditioned place preference. Here, we examined the effects of two nonspecific HDACIs, valproic acid (VPA) and sodium butyrate (NaB), on appetitive learning measured by conditioned stimulus (CS)-induced reinstatement of operant responding. Rats were trained to lever press for food reinforcement and then injected with VPA (50-200 mg/kg, i.p.), NaB (250-1000 mg/kg, i.p.), or saline vehicle (1.0 ml/kg), 2 h before receiving pairings of noncontingent presentation of food pellets preceded by a tone+light cue CS. Rats next underwent extinction of operant responding followed by response-contingent re-exposure to the CS. Rats receiving VPA (100 mg/kg) or NaB (1000 mg/kg) before conditioning displayed significantly higher cue-induced reinstatement than did saline controls. Rats that received either vehicle or VPA (100 mg/kg) before a conditioning session with a randomized relation between presentation of food pellets and the CS failed to show subsequent cue-induced reinstatement with no difference between the two groups. These findings indicate that, under certain contexts, HDACIs strengthen memory formation by specifically increasing the associative strength of the CS, not through an increasing motivation to seek reinforcement.
Assuntos
Condicionamento Clássico/efeitos dos fármacos , Condicionamento Operante/efeitos dos fármacos , Sinais (Psicologia) , Inibidores de Histona Desacetilases/farmacologia , Reforço Psicológico , Análise de Variância , Animais , Butiratos/farmacologia , Relação Dose-Resposta a Droga , Masculino , Ratos , Ratos Sprague-Dawley , Esquema de Reforço , Ácido Valproico/farmacologiaRESUMO
Knowledge of drug concentrations in the brains of behaving subjects remains constrained on a number of dimensions, including poor temporal resolution and lack of real-time data. Here, however, we demonstrate the ability of electrochemical aptamer-based sensors to support seconds-resolved, real-time measurements of drug concentrations in the brains of freely moving rats. Specifically, using such sensors, we achieve <4 µM limits of detection and 10-s resolution in the measurement of procaine in the brains of freely moving rats, permitting the determination of the pharmacokinetics and concentration-behavior relations of the drug with high precision for individual subjects. In parallel, we have used closed-loop feedback-controlled drug delivery to hold intracranial procaine levels constant (±10%) for >1.5 hours. These results demonstrate the utility of such sensors in (i) the determination of the site-specific, seconds-resolved neuropharmacokinetics, (ii) enabling the study of individual subject neuropharmacokinetics and concentration-response relations, and (iii) performing high-precision control over intracranial drug levels.
Assuntos
Encéfalo , Procaína , Ratos , Animais , RetroalimentaçãoRESUMO
Previous studies have shown that brief access to cocaine yields an increase in D2 receptor binding in the medial prefrontal cortex (mPFC), but that extended access to cocaine results in normalized binding of D2 receptors (i.e. the D2 binding returned to control levels). Extended-access conditions have also been shown to produce increased expression of the NR2 subunit of the N-Methyl-D-aspartate receptor in the mPFC. These results implicate disrupted glutamate and dopamine function within this area. Therefore, in the present study, we monitored glutamate and dopamine content within the mPFC during, or 24 hours after, cocaine self-administration in animals that experienced various amounts of exposure to the drug. Naïve subjects showed decreased glutamate and increased dopamine levels within the mPFC during cocaine self-administration. Exposure to seven 1-hour daily cocaine self-administration sessions did not alter the response to self-administered cocaine, but resulted in decreased basal dopamine levels. While exposure to 17 1-hour sessions also resulted in reduced basal dopamine levels, these animals showed increased dopaminergic, but completely diminished glutamatergic, response to self-administered cocaine. Finally, exposure to 17 cocaine self-administration sessions, the last 10 of which being 6-hour sessions, resulted in diminished glutamatergic response to self-administered cocaine and reduced basal glutamate levels within the mPFC while normalizing (i.e. causing a return to control levels) both the dopaminergic response to self-administered cocaine as well as basal dopamine levels within this area. These data demonstrate directly that the transition to escalated cocaine use involves progressive changes in dopamine and glutamate function within the mPFC.
Assuntos
Cocaína/farmacologia , Inibidores da Captação de Dopamina/farmacologia , Dopamina/metabolismo , Ácido Glutâmico/metabolismo , Córtex Pré-Frontal/metabolismo , Animais , Transtornos Relacionados ao Uso de Cocaína/fisiopatologia , Condicionamento Operante , Masculino , Córtex Pré-Frontal/fisiologia , Ratos , Ratos Sprague-Dawley , Esquema de Reforço , AutoadministraçãoRESUMO
The monitoring of specific molecules in the living body has historically required sample removal (e.g., blood draws, microdialysis) followed by analysis via cumbersome, laboratory-bound processes. Those few exceptions to this rule (e.g., glucose, pyruvate, the monoamines) are monitored using "one-off" technologies reliant on the specific enzymatic or redox reactivity of their targets, and thus not generalizable to the measurement of other targets. In response we have developed in vivo electrochemical aptamer-based (E-AB) sensors, a modular, receptor-based measurement technology that is independent of the chemical reactivity of its targets, and thus has the potential to be generalizable to a wide range of analytes. To further the adoption of this in vivo molecular measurement approach by other researchers and to accelerate its ultimate translation to the clinic, we present here our standard protocols for the fabrication and use of intravenous E-AB sensors.
Assuntos
Técnicas Eletroquímicas , Aptâmeros de Nucleotídeos , Técnicas Biossensoriais , OxirreduçãoRESUMO
Estradiol modulates the rewarding and reinforcing properties of cocaine in females, including an increase in selection of cocaine over alternative reinforcers. However, the effects of estradiol on male cocaine self-administration behavior are less studied despite equivalent levels of estradiol in the brains of adult males and females, estradiol effects on motivated behaviors in males that share underlying neural substrates with cocaine reinforcement as well as expression of estrogen receptors in the male brain. Therefore, we sought to characterize the effects of estradiol in males on choice between concurrently-available cocaine and food reinforcement as well as responding for cocaine or food in isolation. Male castrated rats (n=46) were treated daily with estradiol benzoate (EB) (5µg/0.1, S.C.) or vehicle (peanut oil) throughout operant acquisition of cocaine (1mg/kg, IV; FI20 sec) and food (3×45mg; FI20 sec) responding, choice during concurrent access and cocaine and food reinforcement under progressive ratio (PR) schedules. EB increased cocaine choice, both in terms of percent of trials on which cocaine was selected and the proportion of rats exhibiting a cocaine preference as well as increased cocaine, but not food, intake under PR. Additionally, within the EB treated group, cocaine-preferring rats exhibited enhanced acquisition of cocaine, but not food, reinforcement whereas no acquisition differences were observed across preferences in the vehicle treated group. These findings demonstrate that estradiol increases cocaine choice in males similarly to what is observed in females.
Assuntos
Comportamento de Escolha/efeitos dos fármacos , Cocaína/administração & dosagem , Inibidores da Captação de Dopamina/administração & dosagem , Estradiol/análogos & derivados , Animais , Condicionamento Operante/efeitos dos fármacos , Estradiol/farmacologia , Masculino , Ratos , Ratos Sprague-Dawley , Reforço Psicológico , AutoadministraçãoRESUMO
The ability to measure drugs in the body rapidly and in real time would advance both our understanding of pharmacokinetics and our ability to optimally dose and deliver pharmacological therapies. To this end, we are developing electrochemical aptamer-based (E-AB) sensors, a seconds-resolved platform technology that, as critical for performing measurements in vivo, is reagentless, reversible, and selective enough to work when placed directly in bodily fluids. Here we describe the development of an E-AB sensor against irinotecan, a member of the camptothecin family of cancer chemotherapeutics, and its adaptation to in vivo sensing. To achieve this we first re-engineered (via truncation) a previously reported DNA aptamer against the camptothecins to support high-gain E-AB signaling. We then co-deposited the modified aptamer with an unstructured, redox-reporter-modified DNA sequence whose output was independent of target concentration, rendering the sensor's signal gain a sufficiently strong function of square-wave frequency to support kinetic-differential-measurement drift correction. The resultant, 200 µm-diameter, 3 mm-long sensor achieves 20 s-resolved, multi-hour measurements of plasma irinotecan when emplaced in the jugular veins of live rats, thus providing an unprecedentedly high-precision view into the pharmacokinetics of this class of chemotherapeutics.
RESUMO
The electrochemical aptamer-based (E-AB) sensing platform appears to be a convenient (rapid, single-step, and calibration-free) and modular approach to measure concentrations of specific molecules (irrespective of their chemical reactivity) directly in blood and even in situ in the living body. Given these attributes, the platform may thus provide significant opportunities to render therapeutic drug monitoring (the clinical practice in which dosing is adjusted in response to plasma drug measurements) as frequent and convenient as the measurement of blood sugar has become for diabetics. The ability to measure arbitrary molecules in the body in real time could even enable closed-loop feedback control over plasma drug levels in a manner analogous to the recently commercialized controlled blood sugar systems. As initial exploration of this, we describe here the selection of an aptamer against vancomycin, a narrow therapeutic window antibiotic for which therapeutic monitoring is a critical part of the standard of care, and its adaptation into an electrochemical aptamer-based (E-AB) sensor. Using this sensor, we then demonstrate: (i) rapid (seconds) and convenient (single-step and calibration-free) measurement of plasma vancomycin in finger-prick-scale samples of whole blood, (ii) high-precision measurement of subject-specific vancomycin pharmacokinetics (in a rat animal model), and (iii) high-precision, closed-loop feedback control over plasma levels of the drug (in a rat animal model). The ability to not only track (with continuous-glucose-monitor-like measurement frequency and convenience) but also actively control plasma drug levels provides an unprecedented route toward improving therapeutic drug monitoring and, more generally, the personalized, high-precision delivery of pharmacological interventions.
Assuntos
Antibacterianos/sangue , Aptâmeros de Nucleotídeos/química , Monitoramento de Medicamentos/métodos , Técnicas Eletroquímicas/métodos , Vancomicina/sangue , Animais , Antibacterianos/química , Bovinos , Masculino , Ratos Sprague-Dawley , Vancomicina/químicaRESUMO
Similar to the pattern observed in people with substance abuse disorders, laboratory animals will exhibit escalation of cocaine intake when the drug is available over prolonged periods of time. Here, we investigated the contribution of behavioral contingency of cocaine administration on escalation of cocaine intake and gene expression in the dorsal medial prefrontal cortex (dmPFC) in adult male rats. Rats were allowed to self-administer intravenous cocaine (0.25 mg/infusion) under either limited cocaine-(1 h/day), prolonged cocaine-(6 h/day), or limited cocaine-(1 h/day) plus yoked cocaine-access (5 h/day); a control group received access to saline (1 h/day). One day after the final self-administration session, the rats were euthanized and the dmPFC was removed for quantification of mRNA expression of critical glutamatergic signaling genes, Homer2, Grin1, and Dlg4, as these genes and brain region have been previously implicated in addiction, learning, and memory. All groups with cocaine-access showed escalated cocaine intake during the first 10 min of each daily session, and within the first 1 h of cocaine administration. Additionally, the limited-access + yoked group exhibited more non-reinforced lever responses during self-administration sessions than the other groups tested. Lastly, Homer2, Grin1, and Dlg4 mRNA were impacted by both duration and mode of cocaine exposure. Only prolonged-access rats exhibited increases in mRNA expression for Homer2, Grin1, and Dlg4 mRNA. Taken together, these findings indicate that both contingent and non-contingent "excessive" cocaine exposure supports escalation behavior, but the behavioral contingency of cocaine-access has distinct effects on the patterning of operant responsiveness and changes in mRNA expression.
Assuntos
Transtornos Relacionados ao Uso de Cocaína/metabolismo , Cocaína/administração & dosagem , Proteína 4 Homóloga a Disks-Large/biossíntese , Proteínas de Arcabouço Homer/biossíntese , Receptores de N-Metil-D-Aspartato/biossíntese , Animais , Comportamento Aditivo/genética , Comportamento Aditivo/metabolismo , Cocaína/efeitos adversos , Transtornos Relacionados ao Uso de Cocaína/genética , Proteína 4 Homóloga a Disks-Large/genética , Expressão Gênica , Proteínas de Arcabouço Homer/genética , Masculino , Córtex Pré-Frontal/efeitos dos fármacos , Córtex Pré-Frontal/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/genética , AutoadministraçãoRESUMO
Electrochemical, aptamer-based (E-AB) sensors support the continuous, real-time measurement of specific small molecules directly in situ in the living body over the course of many hours. They achieve this by employing binding-induced conformational changes to alter electron transfer from a redox-reporter-modified, electrode-attached aptamer. Previously we have used voltammetry (cyclic, alternating current, and square wave) to monitor this binding-induced change in transfer kinetics indirectly. Here, however, we demonstrate the potential advantages of employing chronoamperometry to measure the change in kinetics directly. In this approach target concentration is reported via changes in the lifetime of the exponential current decay seen when the sensor is subjected to a potential step. Because the lifetime of this decay is independent of its amplitude (e.g., insensitive to variations in the number of aptamer probes on the electrode), chronoamperometrically interrogated E-AB sensors are calibration-free and resistant to drift. Chronoamperometric measurements can also be performed in a few hundred milliseconds, improving the previous few-second time resolution of E-AB sensing by an order of magnitude. To illustrate the potential value of the approach we demonstrate here the calibration-free measurement of the drug tobramycin in situ in the living body with 300 ms time resolution and unprecedented, few-percent precision in the determination of its pharmacokinetic phases.
Assuntos
Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais/métodos , Técnicas Eletroquímicas/métodos , Tobramicina/sangue , Animais , Eletrodos , Transporte de Elétrons , Feminino , Cinética , Masculino , Ratos Sprague-Dawley , Tobramicina/farmacocinéticaRESUMO
Repeated cocaine administration induces many long-term structural and molecular changes in the dorsal medial prefrontal cortex (dmPFC) and are known to underlie aspects of cocaine-seeking behavior. DNA methylation is a key long-lasting epigenetic determinant of gene expression and is implicated in neuroplasticity, however, the extent to which this epigenetic modification is involved in the neuroplasticity associated with drug addiction has received limited attention. Here, we examine the relation between DNA methylation and gene expression within the dorsal medial prefrontal cortex (dmPFC) following limited cocaine self-administration (1â¯h/day), prolonged cocaine self-administration (6â¯h/day), and saline self-administration (1â¯h/day). Rats were fitted with intravenous catheters and allowed to lever press for saline or cocaine (0.25 mg/kg/0.1â¯mL infusion) in the different access conditions for 20 days. Prolonged-access rats exhibited escalation in cocaine intake over the course of training, while limited-access rats did not escalate cocaine intake. Additionally, limited-access and prolonged-access rats exhibited unique Homer2 epigenetic profiles and mRNA expression. In prolonged-access rats, Homer2 mRNA levels in the dmPFC were increased, which was accompanied by decreased DNA methylation and p300 binding within the Homer2 promoter. Limited-access animals exhibited decreased DNA methylation, decreased DNA hydroxymethylation, and increased p300 binding within the Homer2 promoter. These data indicate that distinct epigenetic profiles are induced by limited-versus prolonged-access self-administration conditions that contribute to transcriptional profiles and lend support to the notion that covalent modification of DNA is implicated in addiction-like changes in cocaine-seeking behavior.
Assuntos
Transtornos Relacionados ao Uso de Cocaína/metabolismo , Metilação de DNA/efeitos dos fármacos , Epigênese Genética/efeitos dos fármacos , Proteínas de Arcabouço Homer/metabolismo , Córtex Pré-Frontal/efeitos dos fármacos , Transcriptoma/efeitos dos fármacos , Animais , Cocaína/administração & dosagem , Inibidores da Captação de Dopamina/administração & dosagem , Comportamento de Procura de Droga/fisiologia , Proteína p300 Associada a E1A/metabolismo , Proteínas de Arcabouço Homer/genética , Masculino , Córtex Pré-Frontal/metabolismo , Regiões Promotoras Genéticas , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , AutoadministraçãoRESUMO
By, in effect, rendering pharmacokinetics an experimentally adjustable parameter, the ability to perform feedback-controlled dosing informed by high-frequency in vivo drug measurements would prove a powerful tool for both pharmacological research and clinical practice. Efforts to this end, however, have historically been thwarted by an inability to measure in vivo drug levels in real time and with sufficient convenience and temporal resolution. In response, we describe a closed-loop, feedback-controlled delivery system that uses drug level measurements provided by an in vivo electrochemical aptamer-based (E-AB) sensor to adjust dosing rates every 7 s. The resulting system supports the maintenance of either constant or predefined time-varying plasma drug concentration profiles in live rats over many hours. For researchers, the resultant high-precision control over drug plasma concentrations provides an unprecedented opportunity to (1) map the relationships between pharmacokinetics and clinical outcomes, (2) eliminate inter- and intrasubject metabolic variation as a confounding experimental variable, (3) accurately simulate human pharmacokinetics in animal models, and (4) measure minute-to-minute changes in a drug's pharmacokinetic behavior in response to changing health status, diet, drug-drug interactions, or other intrinsic and external factors. In the clinic, feedback-controlled drug delivery would improve our ability to accurately maintain therapeutic drug levels in the face of large, often unpredictable intra- and interpatient metabolic variation. This, in turn, would improve the efficacy and safety of therapeutic intervention, particularly for the most gravely ill patients, for whom metabolic variability is highest and the margin for therapeutic error is smallest.