Second‐by‐second measurement of acetylcholine release in prefrontal cortex

Abstract

Microdialysis has been widely used to measure acetylcholine (ACh) release in vivo and has provided important insights into the regulation of cholinergic transmission. However, microdialysis can be constrained by limited spatial and temporal resolution. The present experiments utilize a microelectrode array (MEA) to rapidly measure ACh release and clearance in anaesthetized rats. The array electrochemically detects, on a second-by-second basis, changes in current selectively produced by the hydrolysis of ACh to choline (Ch) and the subsequent oxidation of choline and hydrogen peroxidase (H(2)O(2)) at the electrode surface. In vitro calibration of the microelectrode revealed linear responses to ACh (R(2) = 0.9998), limit of detection of 0.08 microm, and signal-to-noise ratio of 3.0. The electrode was unresponsive to ascorbic acid (AA), dopamine (DA), or norepinephrine (NE) interferents. In vivo experiments were conducted in prefrontal cortex (PFC) of anaesthetized rats. Pressure ejections of ACh (10 mm; 40 nL) through an adjoining micropipette produced a rapid rise in current, reaching maximum amplitude in approximately 1.0 s and cleared by 80% within 4-11 s. Endogenously released ACh, following local depolarization with KCl (70 mm; 40, 160 nL), was detected at values as low as 0.05 microm. These signals were volume-dependent and cleared within 4-12 s. Finally, nicotine (1.0 mm, 80 nL) stimulated ACh signals. Nicotine-induced signals reflected the hydrolysis of ACh by endogenous acetylcholinesterase (AChE) as inhibition of the enzyme following perfusion with neostigmine (10 microm) attenuated the signal (40-94%). Collectively, these data validate a novel method for rapidly measuring cholinergic transmission in vivo with a spatial and temporal resolution that far exceeds conventional microdialysis.

A Cholinergic-Dependent Role for the Entorhinal Cortex in Trace Fear Conditioning

“…This contrasts with trace eyeblink conditioning, for which hippocampal-dependent, temporally selective mechanisms have been proposed (Grossberg and Merrill, 1992;Levy et al, 2005;Hasselmo et al, 2007). Nevertheless, acetylcholine is rapidly degraded by hydrolysis in the cortex (Bruno et al, 2006), which may limit the range of acceptable trace intervals. This would be consistent with the facilitation of trace conditioning by cholinesterase inhibitors that has been observed in young rats (Hunt and Richardson, 2007).…”

Abnormal Neurotransmitter Release Underlying Behavioral and Cognitive Disorders: Toward Concepts of Dynamic and Function-Specific Dysregulation

“…Recent advances in the development of new methods for monitoring neurotransmitter release at high temporal and spatial resolution (Venton and Wightman, 2003;Kulagina and Michael, 2003;Burmeister et al, 2000Burmeister et al, , 2002Burmeister et al, , 2006Parikh et al, 2004Bruno et al, 2006a, b;Dale et al, 2005;Day et al, 2006) assist in revealing the multiple and highly dynamic modes of neurotransmitter release. Furthermore, recent experiments provide stark support for the importance of the view that the demonstration and the nature of abnormal neurotransmitter release in animal models of neuropsychiatric disorders requires experiments examining neurotransmitter systems of interest, whereas they are being challenged by relevant behavioral or cognitive functions (references above).…”

Decoupling of Acetylcholine Influx and Theta Power in the Hippocampus

“…The measurement of choline concentration was made possible by coating the amperometry sensors with choline oxidase and a size-exclusion polymer to limit the signal from potential electroactive interferents. Electrons donated from the oxidation of choline through the H 2 O 2 intermediate is detected as a current at the recording site held at a positive potential (Bruno et al, 2006). It is important to note that this method of estimating ACh concentration assumes that all ACh released into CA1 is metabolized to choline by the extremely effective acetylcholine esterase (AChE), but does not take into account possible direct choline releases or concentration of preexisting choline.…”