Cholinergic, noradrenergic, and serotonergic inhibition of fast synaptic transmission in spinal lumbar dorsal horn of rat

doi:10.1016/S0361-9230(01)00470-1

Brain Research Bulletin

Volume 54, Issue 6, April 2001, Pages 639-647

https://doi.org/10.1016/S0361-9230(01)00470-1 Get rights and content

Abstract

It is known that spinal nociceptive sensory transmission receives descending inhibitory and facilitatory modulation from supraspinal structures. Glutamate is the major fast excitatory transmitter between primary afferent fibers and spinal dorsal horn neurons. In whole-cell patch clamp recordings from dorsal horn neurons in spinal slices, we investigated synaptic mechanisms for inhibitory modulation at the lumbar level of the spinal cord. Application of the cholinergic receptor agonist carbachol produced a dose-dependent inhibition of glutamate-mediated excitatory postsynaptic currents (EPSCs) (IC₅₀ 13 μM). Postsynaptic injection of two different types of G-protein inhibitors, guanosine 5′-O-2-thiophosphate or guanosine 5′-O-3-thiotriphosphate, blocked the inhibition produced by carbachol. Clonidine, a selective α-adrenergic receptor agonist, also produced a dose-dependent inhibition of EPSCs (IC₅₀ 7 μM) that was reduced by postsynaptic inhibition of G-proteins. The inhibitory effect of serotonin was likewise mediated by postsynaptic G-proteins. Our results suggest that activation of postsynaptic neurotransmitter receptors plays a critical role in inhibition of glutamate mediated sensory responses by acetylcholine, norepinephrine, and serotonin. Our results support the hypothesis that descending sensory modulation may be mediated by multiple neurotransmitter receptors in the spinal cord.

Introduction

Spinal nociceptive transmission is modulated by an endogenous antinociceptive or analgesic system, consisting of the midbrain periaqueductal gray (PAG) and the rostral ventral medulla (RVM). The RVM serves as an important relay for descending influences from the PAG to the spinal cord 3, 15, 16, 47, 60, 74, 75. Activation of neurons in the RVM inhibits spinal nociceptive transmission and behavioral nociceptive reflexes. The inhibitory effect is mediated directly by descending pathways projecting bilaterally in the dorsolateral funiculi, and indirectly by descending activation of local spinal inhibitory neurons. In the spinal cord, muscarinic, noradrenergic, and serotonergic receptors are important for descending inhibition of behavioral nociceptive reflexes 3, 14, 15, 16, 18, 24, 68, 69, 70, 71, 72, 73, 74, 75.

Electrophysiological studies using intracellular or whole-cell patch-clamp recordings of dorsal horn neurons allow investigation into the cellular mechanisms for the antinociceptive or analgesic effects induced by these transmitters. In anesthetized whole animals, electrical stimulation applied to sites within the nucleus raphe magnus or PAG produced inhibitory postsynaptic potentials (IPSPs) in dorsal horn neurons including ascending projection spinothalamic tract cells 33, 66, 67. More detailed pharmacological analysis came from studies using an in vitro brain/spinal cord slice preparation. In trigeminal nuclei, all three major transmitters, acetylcholine, serotonin, and norepinephrine are reported to inhibit glutamatergic transmission 55, 56. In the lumbar spinal cord, less is known about the synaptic mechanisms underlying sensory inhibition by carbachol, clonidine, and serotonin.

In the present study, we investigated inhibitory modulation of fast excitatory synaptic transmission induced by primary afferent fiber stimulation in the dorsal root entry zone. Glutamate is the major fast excitatory transmitter between primary sensory afferent fibers and dorsal horn neurons 30, 31, 43, 65. Synaptic responses were recorded from neurons in superficial dorsal horn, which are important for receiving sensory information from the periphery 28, 34. Due to a large population of receptor subtypes for each class of transmitter investigated (acetylcholine, serotonin, and norepinephrine), we used pharmacological agents which were used in previous behavioral experiments in vivo 69, 70, 71. Thus, we provide a comparable set of information on the regulation of glutamate-mediated sensory synaptic transmission in the lumbar spinal cord.

Section snippets

Slice preparation

Transverse spinal cord slices from postnatal day 4 to day 21 rats (Sprague-Dawley; Harlan) were prepared. Rats were deeply anesthetized with halothane and then cooled on ice for 10 min. The lumbar spinal cord was exposed, and ice-cold physiological saline solution (all in mM: NaCl, 113; KCl, 3; NaHCO₃, 25; NaH₂PO₃, 1; CaCl₂, 2; MgCl₂, 1; D-glucose, 25; equilibrated with 95% O₂ and 5% CO₂, pH 7.3) was poured over to cool the spinal cord. A segment of lumbar spinal cord was quickly removed and

Inhibition by activation of muscarinic receptors

To test if glutamatergic sensory synaptic transmission in lumber spinal cord may be inhibited by activation of cholinergic receptors, we applied carbachol through bath solution and examined its effect on fast EPSCs. Carbachol produced an inhibitory effect on fast EPSCs (Fig. 1A). The inhibitory effect had a rapid onset and was reversible after the washout of carbachol (see Fig. 1B). Different doses of carbachol were tested and the inhibitory effect of carbachol was dose-dependent (Fig. 3).

Discussion

The present study characterized inhibitory modulation of spinal sensory synaptic transmission mediated by multiple neurotransmitters that were previously implicated in sensory modulation in behavioral experiments with intact animals 61, 69, 70, 71. Consistent with previous findings in the trigeminal nuclei, we found that activation of these neurotransmitter receptors inhibited fast sensory synaptic responses in superficial dorsal horn mediated by glutamate. Our results provide direct evidence

Acknowledgements

We want to thank all members of the Zhuo laboratory for helpful suggestions. This work was supported in part by a grant from the National Institutes of Health (NIDA10833 and NINDS38680).

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Cholinergic, noradrenergic, and serotonergic inhibition of fast synaptic transmission in spinal lumbar dorsal horn of rat

Abstract

Introduction

Section snippets

Slice preparation

Inhibition by activation of muscarinic receptors

Discussion

Acknowledgements

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Brain Res.

Brain Res.

Prog. Brain Res.

Life Sci.

Brain Res.

Brain Res.

Brain Res.

Int. J. Neuropharmacol.

Prog. Neurobiol.

Neuroscience

Brain Res.

Prog. Brain Res.

Eur. J. Pharmacol.

Brain Res.

Neuroscience

Neurosci. Lett.

Brain Res.

Brain Res.

Brain Res.

Brain Res.

Prog. Brain Res.

Eur. J. Pharmacol.

Pain

Brain Res.

Brain Res.

Pain

Neuropharmacology

Muscarinic facilitation of GABA release in substantia gelatinosa of the rat spinal dorsal horn

J. Physiol.

The morphology and distribution of neurons containing choline acetyltransferase in the adult rat spinal cordAn immunocytochemical study

J. Comp. Neurol.

Endogenous pain control systemBrainstem spinal pathways and endorphin circuitry

Annu. Rev. Neurosci.

Tonic descending inhibition of spinal cord neurones driven by joint afferents in normal cats and in cats with an inflamed knee joint

Exp. Brain Res.

Periaqueductal gray stimulation-induced inhibition of nociceptive dorsal horn neurons in rats is associated with the release of norepinephrine, serotonin, and amino acids

J. Pharmacol. Exp. Ther.

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Neurotransmitters in nociceptive modulatory circuits

Annu. Rev. Neurosci.

Brainstem modulation of nociception