In situ temporal detection of dopamine exocytosis from l-dopa-incubated MN9D cells using microelectrode array-integrated biochip

Abstract

Dopamine (DA) is an important neurotransmitter, playing a very important role in many neurological disorders. A microelectrode array-integrated biochip has been represented here as a convenient device for in situ temporal detection of DA exocytosis from dopaminergic cells. The biochip is silicon-based and a 5 × 5 array of Au disk microelectrodes is spaced on the 1 mm center of the silicon plate. MN9D, a mouse mesencephalic dopaminergic cell line, has been grown on the surface of the biochip chamber. DA exocytosis from the chip-grown MN9D cells was detected by using amperometry. With the amperometric detection limit of DA at the biochip microelectrodes ranging from 0.06 to 0.21 μM (S/N = 3), the level of K+-induced DA exocytosis from MN9D cells was undetectable. In contrast, after MN9D cells were incubated with l-dopa, a DA precursor, K+-induced DA exocytosis was temporally detected by amperometry. The K+-induced DA release is concentration-dependent and appears to be saturated at the maximum extracellular DA concentration of 281 ± 137 nM (mean ± S.E.) for 8000 viable MN9D cells, when the extracellular K+ concentration increases to 35 mM. High-performance liquid chromatography demonstrates that the K+-stimulated exocytosis from l-dopa-incubated MN9D cells mainly contains DA, and the weight ratio of DA:NE:l-dopa: serotonin is 1.00:0.28:0.06:0.14. These results suggest that MN9D cell has a typical machinery system of a dopaminergic cell, including l-aromatic acid decarboxylase, vesicular monoamine transporter, voltage-sensitive Na+ channels, and voltage-sensitive calcium channels. © 2005 Elsevier B.V. All rights reserved.