Neurotrophin receptor (p75) in the trigeminal thalamus of the rat: development, response to injury, transient vibrissa-related patterning, and retrograde transport.

We report on the transient, patterned expression of p75 in the ventrobasal (VB) thalamus, the major thalamic relay for somatosensation. We immunostained the brains of developing rats ranging in age from embryonic day (E) 14.5 to postnatal day (PD) 15 with an antibody against p75. To compare p75 expression with the developing synaptic organization within VB, we also immunolocalized the synaptic-vesicle-associated protein, synaptophysin (SYN), on alternate sections. p75-immunoreactivity (IR) was dense and uniform in the ventroposterior medial nucleus (VPM) in the late embryonic and early postnatal periods (E 16.5 to PD 3). In contrast, from PD 4-10, p75-IR in the VPM was patterned, reminiscent of cytochrome-oxidase-stained barreloids, a characteristic feature of the VB in rodents. By PD 14, p75-IR in the VPM was no longer detectable. The ventroposterior lateral nucleus (VPL), in contrast, exhibited no p75-IR. No p75-IR was detected in the ventroposterior lateral nucleus (VPL) at any developmental stage in which VPM could be distinguished from VPL. Light, but clearly patterned SYN-IR, first detectable on PD 2-3, increased in intensity in both VPL and VPM through PD 15. Sectioning the infraorbital nerve on PD 0 resulted in blurred patterns of p75- and SYN-IR within VPM in PD 7-9 rat pups. Removing large portions of the somatosensory cortex on PD 0 resulted in subsequent greatly reduced p75- and SYN-IR within VB. To specify the source of the p75-IR terminals, we stereotaxically injected into the VPM of PD 4-5 rats a monoclonal antibody to p75. One to 2 days later, IR of retrogradely transported p75 antibodies could be traced within axons and cell bodies of neurons associated with the trigeminothalamic pathway through the caudal diencephalon and mesencephalon; labelling was confined to the contralateral trigeminal principal sensory nucleus. The observed, transiently patterned p75-IR in VPM the early postpartum period suggests a role for p75 in synaptogenesis and pattern formation.

Distribution of the low-affinity neurotrophin receptor (p75) in the developing trigeminal brainstem complex in the rat.

The low-affinity neurotrophin receptor (p75) binds all members of the neurotrophin family. In the rat, during the first week postpartum, dense p75-immunoreactivity (IR) is present throughout all components of the trigeminal brainstem complex (TBC), largely associated with primary sensory afferents. Within subnucleus caudalis (SpC) of the TBC, intense p75-IR is present in all laminae at birth. During the second and third postnatal weeks, p75-IR in SpC gradually fades within the deeper laminae, becoming generally restricted in the adult to laminae I and II. Similar declines in p75-IR intensity occur in the subnucleus oralis (SpO); in the SpO in the adult, p75-IR is confined to the dorsalmost portion of SpO. In subnucleus interpolaris, an emerging, vibrissa-related pattern of p75-IR is detectable on PD0 (first 24 hr postpartum), which becomes fully differentiated during PD4-PD7. However, this pattern gradually disappears during the third postnatal week. Ventrally in the nucleus principalis (PrV), a pattern of p75-IR that mirrors the topographical arrangement of the vibrissae is detectable by PD0-PD1, is fully differentiated by the end of the first postnatal week, and persists into adulthood. Perinatal unilateral sectioning of the infraorbital nerve on PD0-PD1, but not as late as PD4, disrupts p75-IR patterning in the adult PrV. Although p75 appears to be associated with primary afferent pattern formation, to determine whether it is essential, we examined mutant mice unable to form functional p75. In the TBC of these knockout mice, examined as adults, patterns of cytochrome oxidase staining (which parallel those of p75-IR) appeared to be normal. In summary, during early development, p75 is widely expressed in the TBC during periods of active synaptogenesis and pattern formation, whereas in the adult, its expression is restricted to association with populations of primary sensory afferents. However, the absence of functional p75 in genetically altered mice does not appear to prevent primary afferent pattern formation.

Synaptophysin immunoreactivity and distributions of calcium-binding proteins highlight the functional organization of the rat's dorsal column nuclei.

The mammalian dorsal column nuclei (DCN) are principally composed of the cuneate (CN) and gracile (GN) nuclei. Data presented here support previously published anatomical and functional evidence that the longitudinal organization of the CN and GN reflect the complex role of the DCN in somatosensory processing. The CN is organized longitudinally into three parts. Within the middle portion of this nucleus, primary afferent projections and cuneothalamic cells are concentrated. Although traditional cytoarchitectonic analyses had failed to detect this tripartite organization in rats, we found evidence for it, with a functional middle region, extending approximately 0.2-0.9 mm caudal to the obex, characterized by precise somatotopy of primary afferent terminations and corresponding somatotopy of cytochrome oxidase (CO) blotches. Additional evidence is presented here consistent with a functionally distinct middle region within the rat's CN: (1) patches of dense synaptophysin (a synaptic-vesical-associated protein)-immunoreactivity (SYN-IR) are limited to the middle CN region, coincident with the dense CO blotches; (2) neurons immunoreactive for the calcium-binding proteins calbindin-D28 (CB), calretinin (CR) and parvalbumin (PV) are concentrated in the middle CN region. Furthermore, in adult rats subjected to perinatal forepaw removal, (1) the patterns of SYN-IR in the middle region of the CN are disrupted, as had previously been shown for the patterns of CO blotches; (2) in contrast, however, distributions of CN cells with PV-, CB- and CR-IR are unaffected. Evidence for a tripartite division in the GN is also presented, based on the distributions of cells with PV-, CB- and CR-IR.

The mutability of low-affinity nerve growth factor receptor (p75NGFR) expression in the rat cuneate nucleus following perinatal injury and adult deafferentations: comparisons with cytochrome oxidase.

In normal adult rats, intense immunostaining for the 75 kDa low-affinity receptor for nerve growth factor and other neurotrophins (p75NGFR) is concentrated in the middle region of the cuneate nucleus (CN), distributed in a blotchy pattern similar to that of cytochrome oxidase (CO) activity. In the adult rats, partial dorsal rhizotomies (centered around the 7th and 8th cervical spinal segments) resulted in the complete disappearance of p75NGFR-like immunoreactivity within the ipsilateral CN, but did not affect the distribution of the CO blotches. Perinatal (postnatal day 1-8) damage to the ipsilateral forepaw and subsequent rearing to adulthood also resulted in significant disruption of the topographical expression of p75NGFR-like immunoreactivity within the CN, as well as--as previously reported--disruption of the CO blotches. Although the patterns of staining in intact adult rats are similar for CO staining and for p75NGFR-like immunoreactivity within the CN, the CO staining appears to be primarily associated with postsynaptic cells, while the p75NGFR-like immunostaining appears to be associated with primary afferent terminals.

The up-regulation of trkA and trkB in dorsal column astrocytes following dorsal rhizotomy.

One week after rat dorsal root fibers were severed, immunoreactivity for the high affinity neurotrophin receptors, trkA and trkB, was up-regulated on glial cells in the spinal cord dorsal column. Immunoreactivity for glial fibrillary acidic protein (GFAP) also increased. In contrast, no changes in immunoreactivity for trkA or trkB, or for GFAP were observed in the brainstem cuneate nucleus, a principal termination site for many of the severed dorsal root fibers.

Enhanced cytochrome-oxidase staining of the cuneate nucleus in the rat reveals a modifiable somatotopic map.

Existing cytochrome oxidase (CO)-staining techniques were modified to enhance sensitivity and contrast in order to examine patterns of CO-activity in the dorsal column nuclei (DCN) of adult Long-Evans rats. Within a rostrocaudally limited region in the middle of the cuneate nucleus (CN) distinctive blotches of intense CO-activity were observed. The CO-staining was maximally differentiated approximately 0.3-0.7 mm caudal to the obex. No CO-blotches were observed anywhere else in the DCN. Transganglionic labelling (WGA-HRP) demonstrated that some of the CO-blotches in the rat CN are related to the terminal projection fields of primary afferents from the skin of the forepaws. The corresponding location of primary afferent termination fields and CO-staining patterns supports a tripartite rostrocaudal division in the rat CN, similar to that described by other investigators in cats, monkeys and raccoons. Comparing the patterns of CO-staining to (1) the cytoarchitecture (Nissl-stained sections), or to (2) the dendritoarchitecture (distribution of microtubule-associated protein 2 (MAP2) or to (3) the organization of retrogradely labelled (WGA-HRP/HRP) cuneothalamic cells, revealed no topographical organization corresponding to the CO-blotches. Postnatal (at least up to 11 days postpartum) forepaw deafferentation or removal disrupted the CO-staining pattern in the CN.

Immunocytochemical localization of the low-affinity nerve growth factor receptor (p75NGFR) in the cuneate nucleus of the rat and its relationship to cytochrome-oxidase activity.

Immunohistochemical staining for the 75-kDa, low-affinity nerve growth factor receptor (p75NGFR), within the cuneate nucleus (CN) of the adult rat revealed that this receptor is concentrated rostrocaudally in the middle CN (approximately 0.2-0.9 mm caudal to the obex), corresponding to that portion of the CN receiving densest projections of cutaneous primary afferent terminals. Furthermore, dense patches of p75NGFR-like immunoreactivity appear to correspond to the 'blotches' of cytochrome-oxidase activity observed in the middle region of the CN. This close correspondence between the localization pattern of p75NGFR in the CN and its functional organization suggests an important role for trophic factors in the CN's development and/or maintenance.

The cuneate nucleus in the rat does have an anatomically distinct middle region.

Recently obtained anatomical evidence supports the division of the rat cuneate nucleus (CN) into three rostrocaudal regions, with the middle region receiving a disproportionately greater share of the primary sensory input. The CN in the rat conforms to the basic rostrocaudal CN pattern described in other mammals, including cat, monkey and raccoon.

Organization of cutaneous primary afferent fibers projecting to the dorsal horn in the rat: WGA-HRP versus B-HRP.

Primary afferent projections from cutaneous afferents in the forelimb and hindlimb digits to the dorsal horn (DH) were examined using 4 tracers: (1) 25% free horseradish peroxidase (HRP), (2) 2.5% wheat-germ agglutinin conjugated to horseradish peroxidase (WGA-HRP), (3) a mixture of 25% free HRP and 2.5% WGA-HRP (WGA-HRP/HRP) or (4) 0.1% HRP conjugated to cholera toxin (B-HRP). The tracer was injected intracutaneously into the digits. Three to 4 days later, the rats were perfused transcardially, transverse sections (60-microns thick) were cut and the HRP was reacted using the tetramethyl benzidine (TMB) method. The location of the label was reconstructed by camera lucida drawings. In rats which received an injection of HRP alone, no label was detected in the DH. Rats injected with WGA-HRP had projection patterns similar to those injected with WGA-HRP/HRP. Patterns of labelling with WGA-HRP differed markedly from those with B-HRP. WGA-HRP labelled cutaneous afferents projecting to Rexed's laminae I-III, with the densest label in lamina II; in contrast, B-HRP labelled cutaneous afferents projecting to laminae II-V, with the densest label in laminae III-IV. These results indicate that, for cutaneous primary afferents projecting to the DH, WGA-HRP and B-HRP labelled different subpopulations of fibers, with the B-HRP-labelled subpopulation biased toward afferents of larger diameter. Rostrocaudally, the extent of the densest fiber projections, whether labelled by WGA-HRP or by B-HRP, was essentially the same, but the extent of the less densely labelled projections was much greater with B-HRP than with WGA-HRP. Comparisons of the projection maps from each of the five digits, using either WGA-HRP or B-HRP, indicated that, as seen in transverse sections through the DH, there was extensive overlapping among the labelled cutaneous afferent fibers from adjacent, or even non-adjacent digits.

Somatotopic organization of the dorsal column nuclei in the rat: transganglionic labelling with B-HRP and WGA-HRP.

To analyze the patterns of cutaneous primary afferent fibers projecting to the dorsal column nuclei in the rat, horseradish peroxidase (HRP)-based tracers were injected intracutaneously into a number of discrete regions of the forelimbs and hindlimbs. Three-4 days following the HRP injections, the rats were perfused transcardially; 60 microns transverse sections were cut, and the HRP was reacted using the tetramethyl benzidine method. Comparisons were made of projections following injections with choleragenoid-conjugated horseradish peroxidase (B-HRP) or with wheat-germ agglutinin conjugated to horseradish peroxidase (WGA-HRP). B-HRP and WGA-HRP produced similar patterns of labelling, but B-HRP produced greater intensity of labelling and slightly larger projection areas. In the cuneate nucleus (CN), HRP labelling of primary afferents from small, delimited regions, e.g., from a portion of the skin of a single digit, appeared to be precisely restricted in rostrocaudally oriented columns, with little or no overlap (in the mediolateral and dorsoventral plane) into adjacent regions. With respect to rostrocaudal organization, a region in the CN containing a dense population of cutaneous primary afferent fibers appeared to be similar to the middle, or cluster, region in cats and in raccoons and the pars rotunda in primates. Projection patterns were very consistent from rat to rat, but their somatotopic organization differed from that suggested by electrophysiological studies: cutaneous afferents from forelimb digit 1 projected near the ventral border of the CN; those from digit 5 projected dorsomedially to those from digit 1; the projections from the remaining digits formed a crescent between the projections from digits 1 and 5. In the gracile nucleus, the organization of cutaneous afferent projections from hindlimb digits was more variable and complex than that found in the CN.

Somatotopic organization of the cuneate nucleus in the rat: transganglionic labelling with WGA-HRP.

A novel somatotopic map of primary cutaneous afferents projecting to the cuneate nucleus in the rat was determined by transganglionic transport of wheat-germ agglutinin conjugated to horseradish peroxidase and free horseradish peroxidase. Intracutaneous injections of tracer into different limited regions of the forelimb resulted in discrete areas of label for each injection site, with little or no overlap into other projection areas. The map of cutaneous projections onto the cuneate nucleus revealed by our anatomical tracing provided much more detail than any previous study in the rat, and demonstrated some significant differences from earlier maps based on electrophysiological recordings.

Computer-assisted three-dimensional reconstructions of [14C]-2-deoxy-D-glucose metabolism in cat lumbosacral spinal cord following cutaneous stimulation of the hindfoot.

We report on computer-assisted three-dimensional reconstruction of spinal cord activity associated with stimulation of the plantar cushion (PC) as revealed by [14C]-2-deoxy-D-glucose (2-DG) serial autoradiographs. Moderate PC stimulation in cats elicits a reflex phasic plantar flexion of the toes. Four cats were chronically spinalized at about T6 under barbiturate anesthesia. Four to 11 days later, the cats were injected (i.v.) with 2-DG (100 microCi/kg) and the PC was electrically stimulated with needle electrodes at 2-5 times threshold for eliciting a reflex. Following stimulation, the spinal cord was processed for autoradiography. Subsequently, autoradiographs, representing approximately 8-18 mm from spinal segments L6-S1, were digitized for computer analysis and 3-D reconstruction. Several strategies of analysis were employed: 1) Three-dimensional volume images were color-coded to represent different levels of functional activity. 2) On the reconstructed volumes, "virtual" sections were made in the horizontal, sagittal, and transverse planes to view regions of 2-DG activity. 3) In addition, we were able to sample different regions within the grey and white matter semi-quantitatively (i.e., pixel intensity) from section to section to reveal differences between ipsi- and contralateral activity, as well as possible variation between sections. These analyses revealed 2-DG activity associated with moderate PC stimulation, not only in the ipsilateral dorsal horn as we had previously demonstrated, but also in both the ipsilateral and contralateral ventral horns, as well as in the intermediate grey matter. The use of novel computer analysis techniques--combined with an unanesthetized preparation--enabled us to demonstrate that the increased metabolic activity in the lumbosacral spinal cord associated with PC stimulation was much more extensive than had heretofore been observed.

Plantar motoneuron columns in the rat.

In the rat, the numbers and locations of motoneurons innervating the short plantar muscles of the hindlimb (supplied by the medial and lateral plantar nerves, as well as a branch of the sural nerve) were determined by using both horseradish peroxidase (HRP) and fluorochromes as retrograde labels. Topographical organization within the plantar motor nucleus was examined by exposing individually the cut ends (encapsulated in low melting-point paraffin) of medial plantar, lateral plantar, and sural nerves to HRP. In addition, double-labeling experiments were conducted in which the medial plantar nerve was labeled with one fluorochrome (either true blue or diamidino yellow) and the lateral plantar nerve with another. The plantar motor pool is located in the extreme dorsolateral portion of the ventral horn, usually concentrated in the fifth lumbar (L5) spinal segment. Labeled motoneurons extended caudally into the sixth lumbar (L6) segment and rostrally into portions of the fourth lumber (L4) segment. Motoneurons of the medial plantar, lateral plantar, and sural nerve have overlapping territories. Sural motoneurons (about 70 cells per side) are generally confined to L5, medial plantar motoneurons (about 180 cells per side) tend to be concentrated in caudal L5 and rostral L6, whereas the lateral plantar motoneurons (about 310 cells per side) extend throughout the entire length of the plantar motor pool. The distribution of motoneuronal cell size is unimodal (mean cross-sectional area = 610 +/- 150 microns2). Cell bodies of plantar motoneurons tend to have similar geometries in all three major planes of sectioning. In all, the combined plantar plus sural nerve population amounts to about 560 motoneurons on each side of the spinal cord. On the basis of these data, and those published by others, the innervation of the small muscles of the foot accounts for about 25% of the motor axons carried by the entire sciatic nerve.

Agonistic behavior of the weakly electric fish, Gnathonemus petersii (Mormyridae, Osteoglossomorpha).

The role played by the electric organ discharge (EOD) in eliciting and maintaining social behavior was assessed. First, a description of the motor acts associated with agonistic encounters between resident and intruder Gnathonemus petersii was provided: The temporal distribution of these acts depended on the aggressiveness of the resident. Second, half of the residents and half of the intruders were rendered electrically "silent" by cutting the nerves to the electric organ. Encounters were staged between pairs of intact fish, pairs of silent fish, and pairs in which one fish was intact and the other silent. Silent residents chased and attacked their opponents less often than the intact fish. Silent and intact residents did not differ in their rates of social probing. Intact intruders elicited higher rates of social probing, and the silent animals elicited more attacks. The effects of the fish's ability to emit EODs and perceive conspecific discharges on different motor acts are discussed.