ReviewImaging of neuronal activity in the gut
Introduction
The enteric nervous system is characterized by its unique ability to mediate reflex behaviour independently of input from the brain or spinal cord, which implies the presence of sensory receptors, primary afferent neurons, interneurons and motor neurons. The functions of the enteric nervous system are multiple and include, at least in part, the control of motor activity, secretion, absorption and blood flow in the gastrointestinal tract [1]. These extensive regulatory activities are made possible by the presence of different types of neurons within the wall of the gastrointestinal tract.
Until recently, enteric neurons have mainly been studied using techniques that are limited to single-cell properties, such as micro-electrode recordings, or only applicable to fixed tissues, such as immunohistochemical staining for chemical coding [1]. The enteric nervous system is functionally arranged in excitatory and inhibitory circuits and the neural control of gastrointestinal functions is determined by the amount of activation within these pathways. Further understanding of the neuronal control of gut functions therefore requires information on activation from multiple neurons simultaneously.
Section snippets
Immunocytochemical imaging of markers of neuronal activation
The initial approach to obtain data on the activation of enteric neuronal circuitry was the immunocytochemical demonstration of markers of previous neuronal activation. The detection of proto-oncogene products such as c-Fos has been used as a marker for changes in neuronal activity [2]; however, this approach is hampered by a number of disadvantages such as the limitation to a single stimulus with subsequent tissue fixation, the lack of quantitative information and the loss of information on
Voltage-sensitive dyes
As changes in membrane potential determine the activity of excitable cells such as enteric neurons, monitoring these changes with voltage-sensitive dyes seems a logical and attractive option. The first generation of these dyes was too slow and had low signal-to-noise ratios. More recently, voltage-sensitive dyes with better dynamic characteristics such as 1-(3-sulfonatopropyl)-4-[beta[2-(di-n-octylamino)-naphthyl]vinyl]pyridinium betaine (Di-8-ANEPPS) have been successfully used to monitor fast
Advantages and disadvantages of different imaging techniques
Inherent to the nature of Ca2+ indicators, these molecules are chelators acting as Ca2+ buffering molecules. Because of their high sensitivity, the dyes can be used at intracellular concentrations unlikely to cause significant Ca2+ buffering. Bleaching is inherent to fluorescent chromophores, but is greatly reduced at moderate illumination power. Autofluorescence of constitutive molecules may interfere with the fluorescence of some shorter wavelength or UV wavelength indicators. Cytotoxicity
Conclusions
Over the past few years, several groups have used voltage-sensitive dyes or calcium-indicator dyes to optically monitor the activity of several enteric neurons simultaneously. Although some of the initial practical problems have been solved, the basic mechanisms underlying these optical signals and their relation to membrane-potential changes needs further investigation. Application of these techniques will provide more insight into neuronal interaction and spread of activation. The potential
References and recommended reading
Papers of particular interest, published within the annual period of review,have been highlighted as:
• of special interest
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