Absence of the p75 neurotrophin receptor alters the pattern of sympathosensory sprouting in the trigeminal ganglia of mice overexpressing nerve growth factor - PubMed (original) (raw)

Absence of the p75 neurotrophin receptor alters the pattern of sympathosensory sprouting in the trigeminal ganglia of mice overexpressing nerve growth factor

G S Walsh et al. J Neurosci. 1999.

Abstract

Sympathetic axons invade the trigeminal ganglia of mice overexpressing nerve growth factor (NGF) (NGF/p75(+/+) mice) and surround sensory neurons having intense NGF immunolabeling; the growth of these axons appears to be directional and specific (). In this investigation, we provide new insight into the neurochemical features and receptor requirements of this sympathosensory sprouting. Using double-antigen immunohistochemistry, we demonstrate that virtually all (98%) trigeminal neurons that exhibit a sympathetic plexus are trk tyrosine kinase receptor (trkA)-positive. In addition, the majority (86%) of those neurons enveloped by sympathetic fibers is also calcitonin gene-related peptide (CGRP)-positive; a smaller number of plexuses (14%) surrounded other somata lacking this neuropeptide. Our results show that sympathosensory interactions form primarily between noradrenergic sympathetic efferents and the trkA/CGRP-expressing sensory somata. To assess the contribution of the p75 neurotrophin receptor (p75(NTR)) in sympathosensory sprouting, a hybrid strain of mice was used that overexpresses NGF but lacks p75(NTR) expression (NGF/p75(-/-) mice). The trigeminal ganglia of NGF/p75(-/-) mice, like those of NGF/p75(+/+) mice, have increased levels of NGF protein and display a concomitant ingrowth of sympathetic axons. In contrast to the precise pattern of sprouting seen in the ganglia of NGF/p75(+/+) mice, sympathetic axons course randomly throughout the ganglionic neuropil of NGF/p75(-/-) mice, forming few perineuronal plexuses. Our results indicate that p75(NTR) is not required to initiate or sustain the growth of sympathetic axons into the NGF-rich trigeminal ganglia but rather plays a role in regulating the directional patterns of axon growth.

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Figures

Fig. 1.

Fig. 1.

A, B, Sections of trigeminal ganglia from NGF/p75+/+ mice double-immunostained for trkA (brown reaction product) and TH (blue reaction product). Virtually all (98%) sympathetic perineuronal plexuses are associated with trkA-positive neurons (arrows). Not all trkA-positive somata, however, exhibit a TH–IR perineuronal plexus.C_–_H, Sections of trigeminal ganglia from NGF/p75+/+ mice double-immunostained for CGRP (blue reaction product) and TH (brown reaction product). The majority (86%) of TH–IR perineuronal plexuses is associated with CGRP–IR sensory neurons. Some somata displaying a TH–IR perineuronal plexus are intensely immunoreactive for CGRP (arrows), whereas others show weak CGRP immunoreactivity (double arrows). A smaller percentage (14%) of TH–IR perineuronal plexuses is associated with somata displaying no detectable CGRP immunoreactivity (large arrowhead). Scale bars, 50 μm.

Fig. 2.

Fig. 2.

Size histograms of the total trigeminal neuron population from C57Bl/6 mice (top panel) and NGF/p75+/+ mice (middle panel), as well as those trigeminal neurons that are surrounded by TH–IR fibers in NGF/p75+/+ mice (bottom panel). Each bin width is 4 μm, and the number assigned to each column represents the middle value for each bin. Comparisons of these cell size distributions reveal that the mean neuron size of C57Bl/6 mice is smaller than that of NGF/p75+/+ mice (p < 0.001), and that TH–IR fibers preferentially envelop large-diameter trigeminal neurons of NGF/p75+/+ mice (p < 0.001).

Fig. 3.

Fig. 3.

Sections of trigeminal ganglia from C57Bl/6, BALB/c, NGF/p75+/+, and NGF/p75−/− mice, stained immunohistochemically for TH. The trigeminal ganglia of C57Bl/6 (A) and BALB/c (B) mice possess TH–IR sympathetic fibers forming perivascular plexuses (arrow), as well as a small population of TH-immunopositive dopaminergic sensory neurons. Many TH–IR fibers are seen in ganglia of NGF/p75+/+mice, projecting to a subset of trigeminal somata and enveloping them in a tight perineuronal plexus of fibers (C,E). TH–IR fibers are also present in ganglia of NGF/p75−/− mice, but the pattern of sprouting appears more random and TH–IR plexuses appear less prominent (D, F). Scale bars:A_–_D, 125 μm; E, F, 50 μm.

Fig. 4.

Fig. 4.

Examples of TH–IR perineuronal plexuses in trigeminal ganglia from NGF/p75+/+ and NGF/p75−/− mice. In NGF/p75+/+animals (A, C, E), several TH–IR fibers often converge on the soma of individual sensory neurons, enveloping the perimeter in a tight, dense plexus of fibers. In NGF/p75−/− mice (B,D, F), TH–IR fibers randomly grow within the neuropil, weaving around the cell bodies of sensory neurons, sometimes in close contact with the somata. However, intense perineuronal plexuses that tightly envelop the somata of NGF/p75−/− ganglia are rarely seen. Scale bar, 25 μm.

Fig. 5.

Fig. 5.

Electron photomicrographs of trigeminal ganglia from NGF/p75+/+ and NGF/p75−/− mice. Sensory somata in NGF/p75+/+ mice (A, B) are surrounded by satellite cell processes closely apposed to their outer cell membrane (S1). Some sensory cell bodies (S2) have many clusters of small-diameter unmyelinated axons (stars) immediately adjacent to their plasma membrane. Occasionally, axonal swellings filled with clear vesicles (Ax) are also seen among the bundles of axons. These unmyelinated axons are ensheathed by nonmyelinating Schwann cells (Sc). Sensory somata in NGF/p75−/−mice (C, D) are likewise surrounded by satellite cell processes closely apposed to their outer cell membrane (S1 and S2). Although small-diameter unmyelinated axons (stars) are also associated with the sensory somata of NGF/p75−/−mice, these fibers are fewer in number, do not occur in clusters, and rarely display axonal swellings filled with clear vesicles. Furthermore, these axons are not ensheathed by nonmyelinating Schwann cells but rather appear to be embedded within the processes of satellite cells. Arrowheads indicate intercellular space between S1 and S2, which is filled with collagen fibers. Scale bars, 1 μm.

Fig. 6.

Fig. 6.

Sections of trigeminal ganglia from C57Bl/6, NGF/p75+/+, and NGF/p75−/−mice, stained immunohistochemically for TH, and a bar graph depicting the density of TH–IR axons in the ganglia of these three mouse genotypes. In the trigeminal fiber tracts of C57Bl/6 mice (A), few if any TH–IR axons are evident. In marked contrast, the trigeminal fiber tracts of both NGF/p75+/+ mice (B) and NGF/p75−/− mice (C) possess numerous TH–IR axons coursing in parallel arrangement with the intrinsic sensory fibers. Note the apparent increase in the number of TH–IR axons in fiber tracts of NGF/p75−/− mice relative to NGF/p75+/+ mice. Scale bar, 100 μm. Quantitation of the percentage area occupied by TH–IR axons in trigeminal fiber tracts (D) confirms an increase in the density of TH–IR axons in the fiber tracts of both NGF/p75+/+ and NGF/p75−/− mice, relative to C57Bl/6 mice (*p < 0.001). The density of TH–IR axons in the fiber tracts of NGF/p75−/−mice is also significantly higher than that in NGF/p75+/+ mice (+p < 0.001). Error bars represent SDs.

Fig. 7.

Fig. 7.

Sections of trigeminal ganglia from C57Bl/6, NGF/p75+/+, and NGF/p75−/−mice, stained immunohistochemically for NGF, and a bar graph depicting the levels of NGF protein in the ganglia of these three mouse genotypes. In C57Bl/6 mice (A), few NGF-positive somata are detected. In contrast, many somata in trigeminal ganglia of NGF/p75+/+ mice (B) display strong immunostaining for NGF. Numerous somata in the trigeminal ganglia of NGF/p75−/− mice (C) also display NGF immunoreactivity, but the intensity of immunostaining is moderate as compared with that in C57Bl/6 and NGF/p75+/+ mice. Scale bar, 50 μm. Mean levels of NGF protein in the trigeminal ganglia of C57Bl/6 (n = 5), NGF/p75+/+(n = 8), and NGF/p75−/−(n = 8) mice were measured by a two-site ELISA (D). This analysis reveals that trigeminal ganglia of NGF/p75+/+ and NGF/p75−/− mice both possess significantly higher levels of NGF protein than trigeminal ganglia of age-matched C57Bl/6 mice (*p < 0.01). The amount of NGF protein in trigeminal ganglia of NGF/p75−/− mice is significantly reduced from that of NGF/p75+/+ mice (+p < 0.001). Error bars represent SDs.

Fig. 8.

Fig. 8.

Sections of trigeminal ganglia from C57Bl/6, NGF/p75+/+, and NGF/p75−/− mice stained immunohistochemically for trkA (A, C, E) and CGRP (B, D, F). In C57Bl/6 mice, numerous somata throughout the trigeminal ganglia possess moderate staining for both trkA (A) and CGRP (B). In comparison, sensory somata in NGF/p75+/+ mice display an increased intensity of immunostaining for both trkA (C) and CGRP (D). NGF/p75−/− mice also display strong immunostaining for trkA (E) and CGRP (F). Immunostaining for trkA and CGRP is evident in small- to medium-sized trigeminal somata of C57Bl/6 mice, whereas immunostaining for trkA and CGRP is seen in small- to large-sized somata of NGF/p75+/+ and NGF/p75−/− mice. Scale bar, 50 μm.

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