Electrochemical Nanosensor for Real-Time Direct Imaging of Nitric

Sep 23, 2011 - Immunohistochemical analysis was carried out to confirm the anatomical reliability of the acquired electrochemical NO image. The real-t...
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Electrochemical Nanosensor for Real-Time Direct Imaging of Nitric Oxide in Living Brain Areum Jo,† Hyunkyung Do,‡ Gil-Ja Jhon,*,‡ Minah Suh,*,†,§ and Youngmi Lee*,‡ †

Department of Biological Science, Sungkyunkwan University, Suwon 440-746, South Korea Department of Chemistry and Nano Science, Ewha Womans University, Seoul, 120-750, South Korea § Graduate Program for Health Science and Technology, Sungkyunkwan University, Suwon 440-746, South Korea ‡

bS Supporting Information ABSTRACT: As gaseous nitric oxide (NO), a critical and multifaceted biomarker, diffuses easily once released, identifying the precise sources of NO release is a challenge. This study developed a new technique for real-time in vivo direct NO imaging by coupling an amperometric NO nanosensor with scanning electrochemical microscopy. This technique provides three-dimensional information of the NO releasing sites in an intact living mouse brain with high sensitivity and spatial resolution. Immunohistochemical analysis was carried out to confirm the anatomical reliability of the acquired electrochemical NO image. The real-time NO imaging results were well matched with the corresponding immunohistochemical analysis of neuronal NO synthase immunoreactive (nNOS-IR) cells, i.e., NO releasing sites in a living brain. The imaged NO local concentrations were confirmed to be closely related to the location in depth, the size of the nNOS-IR cell, and the intensity of nNOS immunoreactivity. This paper demonstrates the first direct electrochemical NO imaging of a living brain.

N

itric oxide (NO), synthesized from L-arginine by the NO synthase (NOS) activity, plays a significant role in shaping the brain function.1 3 As one of its many biological/physiological functions, NO regulates neurovascular coupling by maintaining the cerebrovascular tone and controlling vasodilator responses.4,5 In addition, NO plays an important role as a neurotransmitter for synaptic plasticity and neuronal signaling regulation.2,3,6 On the other hand, the overproduction of NO is related to neurotoxicity in the brain tissue, such as tissue injury, cellular apoptosis, and ischemia.7 In neurodegenerative disorders, NO is released excessively as a result of pro-inflammatory responses. In particular, NOS overexpression along with the formation of nitrotyrosine, a nitrosative stress marker, has been observed in the brains of patients with Alzheimer’s disease and Parkinson’s disease.3,7 Therefore, an effective screening method of NO in the brain may provide extremely useful information on the state of neurodegenerative disease and brain perfusion disorder. Real-time monitoring of NO levels in vivo is not straightforward because of the wide range of NO concentrations (subnanomolar to micromolar),8,9 short half-life (typically