Organotypic cultures (OCs) have been widely used to investigate the midbrain dopaminergic system, but only a few studies focused on the functional properties of dopaminergic neurons and their synaptic inputs from dopaminergic and non-dopaminergic neurons also contained in such cultures. In addition, it is not clear whether the culturing process affects the intrinsic neuronal properties and the expression of specific receptors and transporters. We performed patch-clamp recordings from dopaminergic neurons in mesencephalic-striatal co-cultures obtained from transgenic mice expressing green fluorescent protein (GFP) under the tyrosine hydroxylase promoter. Some (10/44) GFP+ neurons displayed a bursting activity that renders the firing of these cells similar to that of the dopaminergic neurons in vivo. The culturing process reduced the hyperpolarization-activated current (I(h) ) and the expression of D₂ receptors. Downregulation of D₂ receptor mRNA and protein was confirmed with reverse transcriptase polymerase chain reaction and Western blotting. Immunocytochemistry revealed that many synaptic terminals, most likely originating from dopaminergic neurons, co-expressed the dopamine (DA) transporter and the vesicular glutamate transporter-2, suggesting a co-release of DA and glutamate. Interestingly, exogenous DA decreased glutamate release in young cultures [days in vitro (DIV)<20] by acting on pre-synaptic D₂ receptors, while in older cultures (DIV>26) DA increased glutamate release by acting on α-1 adrenoreceptors. The facilitatory effect of DA on glutamatergic transmission to midbrain dopaminergic neurons may be important in conditions when the expression of D₂ receptors is compromised, such as long-term treatment with antipsychotic drugs. Our data show that midbrain OCs at DIV>26 may provide a suitable model of such conditions.
Cucchiaroni, M., Freestone, P., Berretta, N., Viscomi, M. T., Bisicchia, E., Okano, H., Molinari, M., Bernardi, G., Lipski, J., Mercuri, N., Guatteo, E., Properties of dopaminergic neurons in organotypic mesencephalic-striatal co-cultures--evidence for a facilitatory effect of dopamine on the glutamatergic input mediated by α-1 adrenergic receptors., <<EUROPEAN JOURNAL OF NEUROSCIENCE>>, 2011; (33/9): 1622-1636 [http://hdl.handle.net/10807/147322]
Properties of dopaminergic neurons in organotypic mesencephalic-striatal co-cultures--evidence for a facilitatory effect of dopamine on the glutamatergic input mediated by α-1 adrenergic receptors.
Viscomi, M. T.;
2010
Abstract
Organotypic cultures (OCs) have been widely used to investigate the midbrain dopaminergic system, but only a few studies focused on the functional properties of dopaminergic neurons and their synaptic inputs from dopaminergic and non-dopaminergic neurons also contained in such cultures. In addition, it is not clear whether the culturing process affects the intrinsic neuronal properties and the expression of specific receptors and transporters. We performed patch-clamp recordings from dopaminergic neurons in mesencephalic-striatal co-cultures obtained from transgenic mice expressing green fluorescent protein (GFP) under the tyrosine hydroxylase promoter. Some (10/44) GFP+ neurons displayed a bursting activity that renders the firing of these cells similar to that of the dopaminergic neurons in vivo. The culturing process reduced the hyperpolarization-activated current (I(h) ) and the expression of D₂ receptors. Downregulation of D₂ receptor mRNA and protein was confirmed with reverse transcriptase polymerase chain reaction and Western blotting. Immunocytochemistry revealed that many synaptic terminals, most likely originating from dopaminergic neurons, co-expressed the dopamine (DA) transporter and the vesicular glutamate transporter-2, suggesting a co-release of DA and glutamate. Interestingly, exogenous DA decreased glutamate release in young cultures [days in vitro (DIV)<20] by acting on pre-synaptic D₂ receptors, while in older cultures (DIV>26) DA increased glutamate release by acting on α-1 adrenoreceptors. The facilitatory effect of DA on glutamatergic transmission to midbrain dopaminergic neurons may be important in conditions when the expression of D₂ receptors is compromised, such as long-term treatment with antipsychotic drugs. Our data show that midbrain OCs at DIV>26 may provide a suitable model of such conditions.
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