Terminals In Superficial Laminae Research Articles

Microglial Refinement of A-Fiber Projections in the Postnatal Spinal Cord Dorsal Horn Is Required for Normal Maturation of Dynamic Touch.

Sensory systems are shaped in postnatal life by the refinement of synaptic connectivity. In the dorsal horn of the spinal cord, somatosensory circuits undergo postnatal activity-dependent reorganization, including the refinement of primary afferent A-fiber terminals from superficial to deeper spinal dorsal horn laminae which is accompanied by decreases in cutaneous sensitivity. Here, we show in the mouse that microglia, the resident immune cells in the CNS, phagocytose A-fiber terminals in superficial laminae in the first weeks of life. Genetic perturbation of microglial engulfment during the initial postnatal period in either sex prevents the normal process of A-fiber refinement and elimination, resulting in an altered sensitivity of dorsal horn cells to dynamic tactile cutaneous stimulation, and behavioral hypersensitivity to dynamic touch. Thus, functional microglia are necessary for the normal postnatal development of dorsal horn sensory circuits. In the absence of microglial engulfment, superfluous A-fiber projections remain in the dorsal horn, and the balance of sensory connectivity is disrupted, leading to lifelong hypersensitivity to dynamic touch.

Presynaptic low- and high-affinity kainate receptors in nociceptive spinal afferents

Presynaptic ionotropic glutamate receptors are increasingly attributed a role in the modulation of sensory input at the first synapse of dorsal root ganglion (DRG) neurons in the spinal dorsal horn. Central terminals of DRG neurons express AMPA and NMDA receptors whose activation modulates the release of glutamate, the main transmitter at these synapses. Previous work, with an antibody that recognizes all low-affinity kainate receptor subunits (GluR5, 6, 7), provided microscopic evidence of presynaptic kainate receptors in unidentified primary afferent terminals in superficial laminae of the spinal dorsal horn (Hwang SJ, Pagliardini S, Rustioni A, Valtschanoff JG. Presynaptic kainate receptors in primary afferents to the superficial laminae of the rat spinal cord. J Comp Neurol 2001; 436: pp. 275–289). We show here that, although all such subunits may be expressed in these terminals, GluR5 is the subunit most readily detectable at presynaptic sites in sections processed for immunocytochemistry. We also show that the high-affinity kainate receptor subunits KA1 and KA2 are expressed in central terminals of DRG neurons and are co-expressed with low-affinity receptor subunits in the same terminals. Quantitative data show that kainate-expressing DRG neurons are about six times more likely to express the P2X 3 subunit of the purinergic receptor than to express substance P. Thus, nociceptive afferents that express presynaptic kainate receptors are predominantly non-peptidergic, suggesting a role for these receptors in the modulation of neuropathic rather than inflammatory pain.

AMPA Receptor Subunits Underlying Terminals of Fine-Caliber Primary Afferent Fibers

Postembedding immunogold electron microscopy was used to determine the relation of primary afferent terminals in superficial laminae of the spinal dorsal horn with AMPA receptor subunits. Immunogold particles coding for GluR1 and GluR2/3 were concentrated at synaptic sites, between 30 nm outside and 40 nm inside the postsynaptic membrane. Immunopositive synapses displayed round vesicles and asymmetric specializations, characteristic of terminals releasing excitatory neurotransmitters; symmetric synapses, characteristic of terminals releasing inhibitory amino acids, were immunonegative. In superficial laminae, large terminals of two main types at the center of a synaptic glomerulus originate from primary afferents: C1 terminals are mainly endings of unmyelinated afferent fibers; C2 terminals are mainly endings of thinly myelinated afferent fibers. Terminals of both types were presynaptic to AMPA subunits, but in different proportions: C1 terminals were related more to GluR1 than to GluR2/3, whereas the reverse was true for C2 terminals. These results suggest that functional properties of peripheral afferents to the spinal cord may be specified by the density and combination of receptor subunits in the postsynaptic membrane, and raise the possibility that calcium-permeable AMPA channels may play a special role in the mediation of sensory input by unmyelinated fibers.

Synaptic interactions between primary afferent terminals and GABA and nitric oxide-synthesizing neurons in superficial laminae of the rat spinal cord

The superficial laminae (I and II) of the spinal dorsal horn receive small caliber primary afferent fibers responsive to noxious stimulation, and contain local circuit neurons that modulate afferent input. Many of these neurons are GABAergic; about a third of these also synthesize nitric oxide. We identified three main morphological types of primary afferent terminals in superficial laminae after injections of a tracer selective for small caliber afferents into the sciatic nerve of rats. The relative densities of the three types varied through the dorsoventral extent of laminae I and II. Synaptic contacts of each type with GABA- and nitric oxide synthase (NOS)-containing dendrites and axon terminals were determined by preembedding and postembedding immunocytochemistry. Nonglomerular primary afferent terminals, likely to originate from peptidergic unmyelinated fibers, were not seen in synaptic contact with either GABA- or NOS-containing neurons. Primary afferent terminals at the center of type 1 glomeruli (C1) and at the center of type 2 glomeruli (C2) are likely to originate from unmyelinated and small myelinated fibers, respectively. GABAergic terminals contacted more C2 than C1 terminals, suggesting more effective presynaptic inhibition of C2 terminals. Many GABAergic terminals were also positive for NOS, but all GABAergic terminals presynaptic to primary afferent terminals were negative for NOS. Only C2 terminals established frequent synapses with NOS-positive dendrites. (ABSTRACT TRUNCATED AT 250 WORDS)

Amino acid immunocytochemistry of primary afferent terminals in the rat dorsal horn

We combined transganglionic tracing methods with postembedding electron microscopic immunocytochemistry to determine whether identified primary afferent fibers terminating in spinal laminae I-IV may use glutamate and aspartate as neurotransmitters. Sciatic injections of wheat-germ agglutinin conjugated to horseradish peroxidase labeled fine afferent fibers with terminals in laminae I-II of the lumbar spinal cord, whereas injections of the B subunit of cholera toxin conjugated to horseradish peroxidase labeled primary afferent terminals in deeper laminae. Many labeled primary afferent terminals in superficial laminae were involved in glomerular synaptic arrangements; others established nonglomerular contacts. Most glomerular arrangements were clearly immunopositive for glutamate, compared with dendrites, astrocytes, or terminals immunopositive for gamma-aminobutyric acid (GABA). The degree of enrichment varied in labeled terminals of different morphological types. Aspartate was enriched, though to a lesser degree than glutamate, in labeled central terminals of glomeruli in superficial laminae. Labeled primary afferent terminals in laminae III-IV were immunopositive for glutamate, though at lower levels than glomerular terminals in superficial laminae. Aspartate was not enriched in these terminals compared with dendrites, glia, and GABA-positive terminals. These results support a neurotransmitter role for glutamate in primary afferents to the dorsal horn. Quantitative differences in the content of glutamate in identified primary afferent terminals may be related to functional differences. Enrichment of aspartate in terminals in superficial but not deep laminae is compatible with a role for this amino acid in sustained, NMDA-mediated phenomena characteristic of activity in fine caliber afferents.

Changes of spinal substance P, calcitonin gene-related peptide, somatostatin, Met-enkephalin and neurotensin in rats in response to formalin-induced pain

Changes of substance P (SP)-, calcitonin gene-related peptide (CGRP)-, somatostatin (SS)-, Met-enkephalin (Met-Enk)- and neurotensin (NT)- immunoreactive materials on two sides of the lumbar dorsal horn were inspected microscopically and quantified with a computer-assisted image processing system in rats with intact or totally transected spinal cord 2 h after injection of 0.2 ml of 0.5% formalin into the right hindpaw subcutaneously. The results showed that the SP-like immunoreactivity (SP-LI), CGRP-LI, SS-LI, Met-Enk-LI, and NT-LI were significantly higher in fibers and terminals in superficial laminae of the dorsal horn ipsilateral to the formalin injection in both of the experimental groups. It is supposed that the increased contents of these peptides reflect an increased biosynthesis, transport, and release of these peptides in primary afferents and spinal intrinsic neurons in response to the long-lasting inflow of noxious messages, and that these changes seem to be produced even in the condition when the supraspinal effects have been excluded.

Glutamate and substance P coexist in primary afferent terminals in superficial laminae of spinal cord : S. De Biasi and A. Rustioni, Proc. Nat. Acad. Sci. (U.S.A.), 85 (1988) 7820–7824

Glutamate and substance P coexist in primary afferent terminals in superficial laminae of spinal cord : S. De Biasi and A. Rustioni, Proc. Nat. Acad. Sci. (U.S.A.), 85 (1988) 7820–7824