ABSTRACT
The spinal serotoninergic projection from the raphe magnus has been shown to modulate nociceptive inputs, and activation of this projection mediates nicotine-elicited analgesia. Here, we investigate the interactions between cholinergic and serotoninergic systems in the spinal cord, by conducting serotonin [5-hydroxytryptamine (5-HT)] efflux experiments on mouse spinal slices. At least three spinal populations of nicotinic receptors are distinguished that affect 5-HT release. The first could be directly located on serotoninergic terminals, is insensitive to nanomolar concentrations of methyllicaconitine (MLA), and may be subjected to a basal (not maximal) cholinergic tone. The second is tonically and maximally activated by endogenous acetylcholine, insensitive to nanomolar concentrations of MLA, and present on inhibitory neurons. The last is also present on inhibitory neurons but is sensitive to nanomolar concentrations of MLA and not tonically activated by acetylcholine. Multiple nicotinic acetylcholine receptor populations thus differentially exert tonic or not tonic control on 5-HT transmission in the spinal cord. These receptors may be major targets for nicotine effects on antinociception. In addition, the presence of a tonic nicotinic modulation of 5-HT release indicates that endogenous acetylcholine plays a role in the physiological regulation of descending 5-HT pathways to the spinal cord.
Subject(s)
Receptors, Nicotinic/drug effects , Serotonin/physiology , Spinal Cord/drug effects , Synaptic Transmission , Animals , Female , In Vitro Techniques , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Nicotinic Agonists/pharmacology , Nicotinic Antagonists/pharmacology , Receptors, Nicotinic/genetics , Receptors, Nicotinic/physiology , Serotonin/metabolism , Spinal Cord/metabolism , Spinal Cord/physiologyABSTRACT
Knockout mice, in which one or more genes of interest are silenced, provide unique opportunities to analyse diverse aspects of gene function in vivo. In particular, the contribution of the encoded protein(s) in complex behaviours can be assessed. Since the first targeted disruption in 1995 of the gene encoding the beta2-subunit of the nicotinic acetylcholine receptor (nAChR), all but a few of the mammalian nAChR subunits have been disrupted (i.e. alpha7, alpha4, alpha3, alpha9, beta4 and beta3). Recent advances brought by genetically modified mice to our understanding of the endogenous composition and role of nAChRs in the nervous system, and of the diverse pharmacological actions of nicotine regarding learning, analgesia, reinforcement, development and aging in the brain will be discussed.
Subject(s)
Mice, Knockout/genetics , Receptors, Nicotinic/drug effects , Receptors, Nicotinic/genetics , Animals , MiceABSTRACT
The neurons of the locus ceruleus are responsible for most of the noradrenergic innervation in the brain and nicotine potentiates noradrenaline release from their terminals. Here we investigated the diversity and subcellular distribution of nicotinic acetylcholine receptors (nAChRs) in the locus ceruleus both somatically, by combining single-cell reverse transcription-PCR with electrophysiological characterization, and at the level of nerve terminals, by conducting noradrenaline efflux experiments. The proportion of neurons in the locus ceruleus expressing the nicotinic subunit mRNAs varied from 100% (beta2) to 3% (alpha2). Yet, two populations of neurons could be distinguished on the basis of the pattern of expression of nAChR mRNAs and electrophysiological properties. One population (type A) of small cells systematically expressed alpha3 and beta4 mRNAs (and often alpha6, beta3, alpha5, alpha4), and nicotinic agonists elicited large currents with a potency order of cytisine > nicotine. Another population (type B) of cells with large soma did not contain alpha3 and beta4 mRNAs but, systematically, alpha6 and beta3 (and often alpha4) and responded to nicotinic agonists in the order of nicotine > cytisine. The nicotinic modulation of noradrenaline release in the hippocampus displayed an order of potency nicotine > cytisine, suggesting that noradrenergic terminals in the hippocampus originate largely from type B cells of the locus ceruleus. Accordingly, immunocytochemical labeling showed that beta3 is present in hippocampal terminals. The alpha6beta3beta2(alpha4) heterooligomer thus behaves as the main nicotinic regulator of the ceruleo-hippocampal pathway.
Subject(s)
Locus Coeruleus/metabolism , Neurons/metabolism , Neurons/physiology , Receptors, Nicotinic/metabolism , Receptors, Nicotinic/physiology , Adrenergic alpha-Agonists/pharmacology , Alkaloids/pharmacology , Animals , Azocines , Dose-Response Relationship, Drug , Hippocampus/metabolism , Immunohistochemistry , Mice , Norepinephrine/pharmacology , Patch-Clamp Techniques , Quinolizines , RNA, Messenger/metabolism , Rats , Rats, Sprague-Dawley , Receptors, Nicotinic/chemistry , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
Nicotine exerts antinociceptive effects by interacting with one or more of the subtypes of nicotinic acetylcholine receptors (nAChRs) that are present throughout the neuronal pathways that respond to pain. To identify the particular subunits involved in this process, we generated mice lacking the alpha4 subunit of the neuronal nAChR by homologous recombination techniques and studied these together with previously generated mutant mice lacking the beta2 nAChR subunit. Here we show that the homozygous alpha4-/- mice no longer express high-affinity [3H]nicotine and [3H]epibatidine binding sites throughout the brain. In addition, both types of mutant mice display a reduced antinociceptive effect of nicotine on the hot-plate test and diminished sensitivity to nicotine in the tail-flick test. Patch-clamp recordings further reveal that raphe magnus and thalamic neurons no longer respond to nicotine. The alpha4 nAChR subunit, possibly associated with the beta2 nAChR subunit, is therefore crucial for nicotine-elicited antinociception.
