Presynaptic localisation of the nicotinic acetylcholine receptor β2 subunit immunoreactivity in rat nigrostriatal dopaminergic neurones

Abstract

Nicotinic acetylcholine receptors (nAChR) are widely distributed in the central nervous system, where they exert a modulatory influence on synaptic transmission. For the striatum, pharmacological evidence supports the presence of presynaptic alpha3beta2* and alpha4beta2* nAChR that modulate dopamine release from nigrostriatal terminals. The objective of this study was to examine the precise subcellular distribution of the nAChR beta2 subunit in these neurones and its localisation at presynaptic sites. Double immunolabelling with tyrosine hydroxylase (TH) at the confocal level revealed that the cell bodies and axon terminals (synaptosomes) of nigrostriatal neurones were also immunoreactive for the nAChR beta2 subunit. Double-preembedding electron microscopy confirmed that beta2-immunogold labelling was enriched in TH-positive terminals in the dorsal striatum. Quantitative analysis of doubly immunogold-labelled sections in postembedding electron microscopy showed that 86% of TH-positive axonal boutons are also labelled for the nAChR beta2 subunit, whereas 45% of beta2 subunit-immunolabeled boutons do not contain TH. Thus the beta2 subunit is localised within at least two populations of axon terminals in the dorsal striatum. In these structures, 15% of beta2 subunit immunoreactivity was at the plasma membrane but was rarely associated with synapses. These findings are compatible with functional presynaptic beta2-containing nAChR that may be stimulated physiologically by acetylcholine that diffuses from synaptic or nonsynaptic sites of acetylcholine release. These results demonstrate the presynaptic localisation of an nAChR subunit in nigrostriatal dopaminergic neurones, providing morphological evidence for the presynaptic nicotinic modulation of dopamine release.

Cholinergic interneuron characteristics and nicotinic properties in the striatum

“…Whether or not intrinsic striatal neurons express ␤2* nAChRs is controversial. Hill et al (1993) detected ␤2-subunit immunoreactivity on cholinergic interneurons, but Jones et al (2001) did not find ␤2-subunit immunoreactive cells in the striatum. Electrophysiologic data, however, support the presence of nAChRs on GABAergic interneurons in the striatum, but not on the medium spiny neurons or cholinergic interneurons.…”

“…Consistent with those results, mutant mice lacking the ␤2 subunit release much less DA in response to an electrical shock delivered in the striatal slice preparation. Other evidence further indicates that striatal DA terminals express ␤2* nAChRs (Jones et al, 2001). It is likely that much of the ACh-dependent modulation of DA release arises from ␤2* nAChRs located on DA terminals.…”

“…Genetic deletion of the ␤2 subunit results in a dramatic decrease or loss of nicotine-evoked effects in the midbrain DA areas and the striatum (Picciotto et al, 1998;Zhou et al, 2001). All DA neurons and many GABA neurons in the VTA and SN express ␤2* nAChRs (Picciotto et al, 1998;Jones et al, 2001;Klink et al, 2001). Activation of these somatic nAChRs increases the excitability, action potential firing, and Ca 2ϩ influx in these neurons (Pidoplichko et al, 1997;Picciotto et al, 1998;Klink et al, 2001).…”

“…Histochemical and ion flux measurements with synaptosomes have indicated that nAChRs are expressed on DA axon terminals in the striatum (Schwartz et al, 1984;Hill et al, 1993;Colquhoun and Patrick, 1997;Wonnacott, 1997;Jones et al, 2001). Electron microscopy applied with immunolabeling for tyrosine hydroxylase and for the nAChR ␤2 subunit revealed that striatal DA axonal boutons and midbrain DA cell bodies express ␤2* nAChRs (Jones et al, 2001).…”

“…Electron microscopy applied with immunolabeling for tyrosine hydroxylase and for the nAChR ␤2 subunit revealed that striatal DA axonal boutons and midbrain DA cell bodies express ␤2* nAChRs (Jones et al, 2001). Whether or not intrinsic striatal neurons express ␤2* nAChRs is controversial.…”