Microdialysis perfusion

Portas, C. M., Thakkar, M., Rainnie, D. McCarley, R. W. (1996). Microdialysis perfusion of 8-hydroxy-2-(di-n-propylamino)tetrahn (8-OH-DPAT) in the dorsal raphe nucleus decreases serotonin release and increases rapid eye movement sleep in the freely moving cat. J. Neurosci. 16, 2820-8. 

Figure 6.4 Microdialysis perfusion of histamine into the cholinergic zone of the basal forebrain increases wakefulness in a dose-dependent fashion. The animals spent almost half of their time in wakefulness during the 2 h period of perfusion of the highest dose (1,000 p,M). In contrast, approximately 12% of the time was spent in wakefulness during the 2 h perfusion period of artificial CSF (i.e. the control day). Adapted from Ramesh et al. (2004).

Thakkar, M. M., Ramesh, V., Strecker, R. E. McCarley, R. W. (2001). Microdialysis perfusion of orexin-A in the basal forebrain increases wakefulness in freely behaving rats. Arch. Ital. Biol. 139, 313-28. 

Sorensen, E., Gronli, J., Bjorvatn, B., Bjorkum, A. Ursin, R. (2001). Sleep and waking following microdialysis perfusion of the selective 5-HTia receptor antagonist p-MPPI into the dorsal raphe nucleus in the freely moving rat. Brain Res. 897, 122 30. 

Moghaddam, B. and Bunney, B.S., Ionic composition of microdialysis perfusing solution alters the pharmacological responsiveness and basal outflow of striatal dopamine, /. Neurochem., 53, 652, 1989. 

McNay EC, Sherwin RS. 2004. From artificial cerebro-spinal fluid (acsf) to artificial extracellular fluid (aecf) microdialysis perfusate composition effects on in vivo brain ecf glucose measurements. J Neurosci Methods 132(1) 35-43. 

Brock, John W., Jeffrey P. Ng, and Joseph B. Justice Jr. 1990. "Effect of Chronic Cocaine on Dopamine Synthesis in the Nucleus Accumbens as Determined by Microdialysis Perfusion with NSD-10." Neuroscience Letters 117 234-39. Broderick, Patricia A. 1985. "In Vivo Electrochemical Studies of Rat Striatal Dopamine and Serotonin Release After Morphine." Life Sciences 36 2269-75. 

Jelev A, Sood S, Liu H, Nolan P, Homer RL (2001) Microdialysis perfusion of 5-HT into hypoglossal motor nucleus differentially modulates genioglossus activity across natural sleep-wake states in rats. J Physiol 532(Pt 2) 467 181... 

Besides the basic apparatus for microdialysis perfusions, fraction collection, and HPLC analysis, several additional instruments and devices are needed, depending on where the microdialysis probe is to be implanted. The most complicated instrumental setup is probably that required for brain dialysis. A stereotaxtic instrument and a stereomicroscope are necessary for precise positioning of microdialysis cannulae into various brain structures. Inhalation anesthesia is preferable and more convenient than injections. However, this type of anesthesia calls for additional equipment, such as air lines, valves, and mixing chamber for halothane or other anesthetic gases, as well as good ventilation of the operation theater. 

The system has been improved through redesigning the closed-flow microdialysis perfusion system of the sensor to minimize the risk of enzyme leakage by immobilizing the enzymes GOD and catalase in an enzyme reactor. 

This chapter describes two microbore LC-based methods for measurement of monoamine neurotransmitters in rat brain dialysates a method for the determination of the catecholamines (CA) noradrenaline (NAD), adrenaline (AD), and dopamine (DA) and a method for the determination of the indoleamine serotonin (5HT) and its major metabolite 5-hydroxyindoleacetic acid (SHIAA) (3). For all compounds, a limit of detection of 1 pmol or less (in an injected volume of 10 pL) can be achieved. Basal serotonin can be measured in microdialysates without including a reuptake inhibitor in the microdialysis perfusion fluid. Both of these systems are routinely used in the authors laboratory for the analysis of dialysates from striatum, hippocampus, and substantia nigra. Both methods are selective, robust, and can be automated. 

Figure 4,7 The effect of perfusion of the microdialysis probe with a medium containing a depolarising (80 mM) concentration of K" ", or Ca +-free medium, for the periods indicated by the bars. The graph shows efflux of noradrenaline in the frontal cortex of anaesthetised rats. Increasing the concentration of K" " in the medium infused via the probe increases noradrenaline efflux whereas removing Ca reduces it...

The ability of the striatum to apparently function normally until it has lost much of its DA can be ascribed in part to denervation supersensitivity, the degeneration of the DA input resulting in an increase in postsynaptic DA receptors and partly to the remaining neurons producing more DA. This is supported by measurements in humans which show that the HVA DA ratio, a measure of DA turnover, is much greater in Parkinsonism patients and by microdialysis in rats with 6-OHDA lesions of the nigrostriatal tract, when the reduction in perfusate (released) DA is very much less than that of neuronal (stored) DA. 

Parsons, L.H., Justice, J.B. Perfusate serotonin increases extracellular dopamine in the nucleus accum-bens as measured by in vivo microdialysis. Brain Res. 606 195, 1993. 

The technique consists of a microdialysis probe, a thin hollow tube made of a semi-permeable membrane usually around 200-500 /xm in diameter, which is implanted into the skin and perfused with a receiver solution that recovers the unbound permeant from the local area. In principle, the driving force of dialysis is the concentration gradient existing between two compartments separated by a semi-permeable membrane. For skin under in vivo conditions, these compartments represent the dermal or subcutaneous extracellular fluid (depending on the probe position) and an artificial physiological solution inside the probe [36-38],... 

The relative recovery (RR) of the probe, essential for data interpretation, is calculated using the retrodialysis method, which assumes that the net transport through the microdialysis membrane from the perfusate to the surrounding tissues equals the net transport from the tissues into the perfusate. The equation for calculation is represented as follows [39] ... 

In vivo microdialysis is based on the principle of dialysis, the process whereby concentration gradients drive the movement of small molecules and water through a semipermeable membrane. In vivo microdialysis involves the insertion of a small semipermeable membrane into a specific region of a living animal, such as the brain. The assembly that contains this semipermeable membrane is called a probe, which is composed of an inlet and an outlet compartment surrounded by a semipermeable membrane (see O Figure 9-1). Using a microinfusion pump set at a low flow rate (0.2-3 /rL/min), an aqueous solution known as the perfusate is pumped into the inlet compartment of the microdialysis probe. Ideally, the... 

Graph used to calculate the point of no net flux for dopamine (DA). Using regression analysis, the extracellular concentration of DA is estimated via the difference method [the DA concentration in the perfusate minus the concentration of DA in the dialysate] plotted against the DA concentration in the perfusate. Values above the zero on the y-axis indicate diffusion to the brain, whereas values below the zero indicate diffusion from the brain. The zero point on the y-axis represents a steady state, at which no net flux of DA occurs across the dialysis membrane and represents the extracellular concentration of DA on the x-axis. Figure from Parsons, L.H., Justice, J.B., Jr. (1994). Quantitative approaches to in vivo brain microdialysis. Crit Rev Neurobiol. 8(3) 189-220... 

After the completion of the in vivo microdialysis experiment, rats are euthanized with an overdose of sodium pentobarbital (100 mg/kg) and perfused intracardially with 0.9% saline followed by 10% formalin. The brains are extracted and stored in a 10% formalin solution until they are sectioned into slices (40-60 mm) consecutively through the guide cannula tract. The sections are mounted onto gelatin-coated glass slides and stained with thionin. An observer unaware of the rat s treatment or results verifies the cannula location for each rat. 

Ogasawara M, Nakajima W, Ishida A, Takada G. 1999. Striatal perfusion of indomethacin attenuates dopamine increase in immature rat brain e3q>osed to anoxia an in vivo microdialysis study. Brain Res 842(2) 487-490. 

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