Introduction
Microdialysis and voltammetry are widely used for monitoring extracellular dopamine in the brain of living animals. It is important to understand the relationship between experimental results and the in vivo dopamine concentration. Calibration methods that permit these relationships to be experimentally determined have not yet been reported. As an alternative to calibration, we have directly compared results obtained simultaneously by voltammetry and microdialysis under identical conditions in vivo.

Methods
Voltammetric microelectrodes (7 mm in diameter, 400-800 mm in length) were placed far away (1mm) from, immediately adjacent to, and at the outlet of microdialysis probes implanted in the striata of chloral hydrate-anesthetized rats. Fast scan cyclic voltammetry was performed simultaneously at all three microelectrodes at 300 V/s with a 200 ms scan interval. Voltammetric recording was performed during electrical stimulation of MFB (45 Hz, 25s, 50 mA rms). Nomifensine (20 mg/kg), cocaine (30 mg/kg), L-dopa (250 mg/kg, given 30 min after 150 mg/kg carbidopa), pargyline (75 mg/kg) and sulpiride (100 mg/kg) were administrated (i.p.) to examine their effect on the stimulation response at all three microelectrodes.

Results and Discussion
Even when the electrically evoked response at the electrode far away from the microdialysis probe was large, there was no detectable response at the electrodes adjacent to and at the outlet of the probe (Figure 1). Administration of nomifensine increased the response at the electrode far from the probe but, on a proportional basis, increased the response at the adjacent and outlet electrodes to a much larger extent (Figure 1, top row). After administration of L-dopa, the response at the electrode far from the probe was significantly increased but the response at the adjacent and outlet electrodes remained undetectable (Figure 1, bottom row).

These results show that uptake prevents diffusion of dopamine to implanted microdialysis probes to a significantly greater extent than it prevents diffusion of dopamine to carbon fiber microelectrodes. Furthermore, the results show that, as a consequence of uptake, the microdialysis recovery of dopamine in these experiments is initially near zero and increases upon uptake inhibition (Figure 1, top row). The apparent increase in recovery after uptake inhibition is opposite to the reported decrease in extraction fraction following uptake inhibition,¹ which suggests that recovery and extraction of dopamine during in vivo experiments are not equivalent.

The no-net-flux and extrapolation to zero flow methods of quantification require equal values of the extraction fraction and relative recovery. Hence, this direct comparison between voltammetry and microdialysis results obtained under identical in vivo conditions leads to the conclusion that these methods may not be appropriate for quantification of dopamine. In particular, since these results suggest that the recovery of dopamine in the absence of uptake inhibition is very small, this work suggests that the no-net-flux method underestimates the basal extracellular concentration of dopamine.

Figure 1. Representative responses to electrical stimulation of the MFB recorded before (‘pre’) and after (‘post’) nomifensine (upper panel) or L-dopa (Lower panel) administration.

References

1. Smith A.D. and Justice J.B. Jr., (1994) The effect of inhibition of synthesis, release, metabolism and uptake on the microdialysis extraction fraction of dopamine. J. Neurosci. Methods 54: 75-82.