SLAS

Microdialysis: An introduction

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Adapted in part with from "Principle of Microdialysis  by Jan Kehr,  Pronexus Analytical AB, with permission.

Microdialysis sampling is a diffusion-based separation method that allows analytes to freely diffuse across a hollow fiber semi-permeable dialysis membrane. This sampling technique has been widely used for in vivo chemical collection.  Microdialysis uses the principle of the dialysis (Greek: separate), in which a membrane (i.e. a microdialysis probe), permeable to water and small solutes, is introduced into an analyte-rich environment, such as living tissue.  The membrane is perfused with a liquid (perfusate) which equilibrates with the fluid outside the membrane by diffusion in both directions.  Osmotic pressure is the driving force. 

History

Originally used to sample from the brain tissue, microdialysis has become a very common method to sample free drug concentrations from any tissue and so a very important tool to determine the pharmacokinetics in these tissues.  Microdialysis in combination with a suitable detection technique allows monitoring of time-dependent changes in local tissue chemistry, for example neurotransmitter release and reuptake, drug delivery or energy metabolism in a particular brain area.  In 1966 Bito et al.[1] described the possibility of using a semi-permeable membrane to sample free amino acids and other electrolytes in the extracellular fluid of brain and blood plasma of the dog. 

In 1972 Delgado et al.[2] reported a construction of a ”dialytrode” for monkeys, which was basically a push-pull cannula with a small (5 x 1 mm) polysulfone membrane bag glued on its tip.  The authors described some conceptual experiments, derived from the established protocols for push-pull experiments: infusing a compounds or labelled precursors into the brain and correlating these effects to brain electrical activity or to a degree of newly synthesised labelled compounds and sampling and subsequent analysis of endogenous compounds such as amino acids. 

It was Ungerstedt and Pycock (1974)[3] who finally succeeded to measure amphetamine-induced release of dopamine-like radioactivity after local prelabeling of brain tissue with tritiated dopamine perfused through a hollow fibre dialysis probe implanted into the rat striatum.

A rapid advance in development of highly sensitive HPLC analytical techniques and technologies of manufacturing small-bore dialysis tubing during the coming years accelerated the research on applications of single fibre dialysis - microdialysis - in neurobiology, pharmacology and physiology from animals (Ungerstedt et al 1982; Hamberger et al 1982; Ungerstedt 1984) to humans (Meyerson et al 1990; Hillered et al 1990; Ungerstedt 1991). Several books on in vivo monitoring include chapters on microdialysis, and a monograph ”Microdialysis in the Neurosciences” was edited by Robinson and Justice in 1991. A series of meetings with emphasis on electrochemical detection and in vivo methods in neuropharmacology was initiated by Prof. C. Marsden in 1982. Gradually, the meetings grew into a form of medium-sized international conferences on "Monitoring Molecules in Neuroscience", which are organized every second year. The proceedings of the conferences can provide a fast overview on major applications areas and technological advancements of in vivo monitoring techniques.

Adapted from Kehr J (1999) Monitoring chemistry of brain microenvironment: biosensors, microdialysis and related techniques, Chapter 41. In: Modern techniques in neuroscience research (Eds U Windhorst and H Johansson) Springer-Verlag GmbH, Heidelberg, 1149-1198.


Image:Microdialysis_Fig_2.gif
Figure1: The first successful demonstration of feasibility to use brain dialysis for measuring neurotransmitter release in vivo. Measuring amphetamine-induced release of DA-like radioactivity after local pre-labeling of the rat striatum with tritiated DA.  Adopted from: Ungerstedt U, Pycock C (1974) Bull Schweiz Akad Med Wiss, 1278:1-13.


Current practice

Microdialysis is a minimally invasive technique allowing in vivo sampling of molecules transported into, or generated within the extracellular space of principally any tissue or organ in the body and also for sampling the body fluids such as blood or CSF. Microdialysis in combination with a suitable detection technique allows monitoring of time-dependent changes in local tissue chemistry, for example neurotransmitter release and reuptake, drug delivery or energy metabolism in a particular brain area. The unsurpassed feature of in vivo microdialysis is its capability to provide information on basal, non-stimulated levels of extracellular neurotransmitters, as well as, on pharmacologically or physiologically stimulated release. This offers a unique opportunity to examine the role of various receptor subtypes in tonic and phasic regulation of neurotransmitter release and metabolism in neuroanatomically relevant brain structures.

Microdialysis technique provides the most comprehensive information on dynamic changes of molecules involved in intercellular communication and metabolism at an integrative (whole body) level, preserving the overall physiological and behavioral functions.

Several independent techniques have demonstrated that the molecular movement within the extracellular space is driven predominantly by diffusion. In an analogous way, the driving force of microdialysis sampling is the diffusion of molecules across the concentration gradients existing between the two compartments separated by the membrane: the tissue/extracellular space compartment and the perfusion fluid inside the probe as the second compartment. Thus, the molecules can move in both directions, which allows simultaneous recovery of endogenous compounds released into the brain microenvironment and at the same time, drugs can be delivered locally through the probe into the sampled area.

Microdialysis can be used for monitoring the kinetics of drug distribution and clearance in different organs, which for the brain studies is of particular interest since it allows evaluating the ability of drugs to penetrate through the blood-brain barrier.


Image:Microdialysis_Fig_1.jpg
Figure 2. The principle of microdialysis sampling and the origin of molecules released into the brain microenvironment. The molecular movement of substances present in the extracellular fluid is driven predominantly by diffusion.

Depending on the concentration gradients across the membrane of the microdialysis probe, the molecules can move in both directions, which allows simultaneous recovery of endogenous compounds and at the same time, drugs can be added to the perfusion medium and delivered locally into the same brain structure.

The most interesting and thoroughly studied molecules are (Fig. 2: depicted by numbers): 1,2,3 - neurotransmitters, neuromodulators and neuropeptides; 3,4 - the neuron-glia interactions, glutamate and GABA, large molecules such as interleukins and trophic factors; 5 - second messengers cAMP, cGMP or arachidonic acid metabolites; 6 - molecules transported from blood capillaries - glucose, nutrients, drugs; 7 - neuro-vascular communication - NO; 8 - molecules transported from or into the CSF.
Adopted from: Kehr J, Yoshitake T (2006) Monitoring brain chemical signals by microdialysis. In: Encyclopedia of Sensors (Eds CA Grimes, EC Dickey and MV Pishko) American Scientific Publishers, USA, pp 287-312.

Current analytical techniques used in conjunction with microdialysis include High Performance Liquid Chromatography (HPLC), gas chromatography, electrophoresis, radioimmunoassays (RIA), enzyme methodologys, LC-MS and LC-MS/MS.

Microtechnology

Due to the small sample dialysate volumes involved, microdialysis is well matched with micro-scale analytical techniques that have evolved with the development of microfluidics and lab-on-a-chip technologies. Figure 3 illustrates one such example.  It comprises a dual electrochemically actuated pump for sensor calibration, an amperometric sensor (2) for e.g. glucose or lactate measurement, a potentiometric sensor (1) for e.g. Na+, K+ or Li+ measurements, a reference electrode (RE) and a needle shaped microdialysis probe. The complete system is realized in a stack of a glass part and a silicon part with total dimensions of about 2.7cm x 1.5cm x 0.1cm.

Image:Microdialysis_Fig_3.jpg
Figure 3: Lay out of the complete µTAS chip, including a microdialysis probe, two sensors, a reference electrode and a dual electrochemically actuated dosing pump for sensor calibration.  Ad Sprenkels, Dorota Pijanowska, Heiko van der Linden, Wouter Olthuis, Piet Bergveld, University of Twente, the Netherlands.

Such technology makes it possible to consider chip-scale microdialysis / analysis systems small enough to be attached to the monitored organism, untetherd to other external devices. 

References

  1. Bito L, Davson H, Levin E, Murray M, Snider N (1966) The concentrations of free amino acids and other electrolytes in cerebrospinal fluid, in vivo dialysate of brain, and blood plasma of the dog. J Neurochem, 13:1057-1067.
  2. Delgado JM, DeFeudis FV, Roth RH, Ryugo DK, Mitruka BM (1972) Dialytrode for long term intracerebral perfusion in awake monkeys. Arch Int Pharmacodyn, 198:9-21.
  3. Hamberger A, Jacobson I, Molin S-O, Nyström B, Sandberg M, Ungerstedt U (1982) Metabolic and transmitter compartments for glutamate. In: (Ed H Bradford) Neurotransmitter interaction and compartmentation. Plenum, New York, pp 359-378

Additional reading

  • Kehr J (1999) Monitoring chemistry of brain microenvironment: biosensors, microdialysis and related techniques, Chapter 41. In: Modern techniques in neuroscience research (Eds U Windhorst and H Johansson) Springer-Verlag GmbH, Heidelberg, 1149-1198.
  • Kehr J, Yoshitake T (2006) Monitoring brain chemical signals by microdialysis. In: Encyclopedia of Sensors, Vol. 6. (Eds CA Grimes, EC Dickey and MV Pishko) American Scientific Publishers, USA, pp 287-312.
  • Kehr J (2007) New methodological aspects of microdialysis. In: Handbook of microdialysis: Methods, Applications and Perspectives (Eds BHC Westerink and T Cremers) Elsevier, The Netherlands, pp 111-129.
  • Hillered L, Persson L, Ponten U, Ungerstedt U (1990) Neurometabolic monitoring of the ischaemic human brain using microdialysis. Acta Neurochirurgica, 102:91-97.
  • Meyerson BA, Linderoth B, Karlsson H, Ungerstedt U (1990) Microdialysis in the human brain: extracellular measurements in the thalamus of parkinsonian patients. Life Sci, 46:301-308.
  • Robinson TE, Justice JB Jr (1991) Microdialysis in the Neurosciences, Techniques in the Behavioral and Neural Sciences Vol. 7, Elsevier, Amsterdam.
  • Roth RH, Allikmets L, Delgado JM (1969) Synthesis and release of noradrenaline and dopamine from discrete regions of monkey brain. Arch Int Pharmacodyn, 181:273-282.
  • Ungerstedt U (1984) Measurement of neurotransmitter release by intracranial dialysis. In: (Ed CA Marsden) Measurement of neurotransmitter release in vivo. Methods in neurosciences, vol 6. Wiley, New York, pp 81-105.
  • Ungerstedt U (1991) Microdialysis--principles and applications for studies in animals and man. J Int Med, 230:365-373.
  • Ungerstedt U, Herrera-Marschitz M, Jungnelius U, Ståhle L, Tossman U, Zetterström T (1982) Dopamine synaptic mechanisms reflected in studies combining behavioural recordings and brain dialysis. In: (Eds M Kotisaka, T Shomori, T Tsukada, GM Woodruff) Advances in dopamine research. Pergamon Press, New York, pp 219-231.
  • Ungerstedt U, Pycock C (1974) Functional correlates of dopamine neurotransmission. Bull Schweiz Akad Med Wis, 30:44-55.

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