L4 Neurons Research Articles

Coding of locomotor phase in populations of neurons in rostral and caudal segments of the neonatal rat lumbar spinal cord.

Several experiments have demonstrated that rostral segments of the vertebrate lumbar spinal cord produce a rhythmic motor output more readily and of better quality than caudal segments. Here we examine how this rostrocaudal gradient of rhythmogenic capability is reflected in the spike activity of neurons in the rostral (L(2)) and caudal (L(5)) lumbar spinal cord of the neonatal rat. The spike activity of interneurons in the ventromedial cord, a region necessary for the production of locomotion, was recorded intracellularly with patch electrodes and extracellularly with tetrodes during pharmacologically induced locomotion. Both L(2) and L(5) neurons tended to be active in phase with their homologous ventral root. L(5) neurons, however, had a wider distribution of their preferred phases of activity throughout the locomotor cycle than L(2) neurons. The strength of modulation of the activity of individual L(2) neurons was also larger than that of L(5) neurons. These differences resulted in a stronger rhythmic signal from the L(2) neuronal population than from the L(5) population. These results demonstrate that the rhythmogenic capability of each spinal segment was reflected in the activity of interneurons located in the same segment. In addition to paralleling the rostrocaudal gradient of rhythmogenic capability, these results further suggest a colocalization of motoneurons and their associated interneurons involved in the production of locomotion.

Reduction in potassium currents in identified cutaneous afferent dorsal root ganglion neurons after axotomy.

Potassium currents have an important role in modulating neuronal excitability. We have investigated the effects of axotomy on three voltage-activated K(+) currents, one sustained and two transient, in cutaneous afferent dorsal root ganglion (DRG) neurons. Fourteen to 21 days after axotomy, L(4) and L(5) DRG neurons were acutely dissociated and were studied 2-8 h after plating. Whole cell patch-clamp recordings were obtained from identified cutaneous afferent neurons (46-50 microm diam); K(+) currents were isolated by blocking Na(+) and Ca(2+) currents with appropriate ion replacement and channel blockers. Separation of the current components was achieved on the basis of sensitivity to dendrotoxin or 4-aminopyridine and by the response to variation in conditioning voltage. Both control and injured neurons displayed qualitatively similar complex K(+) currents composed of distinct kinetic and pharmacological components. Three distinct K(+) current components, a sustained (I(K)) and two transient (I(A) and I(D)), were identified in variable proportions. However, total peak current was reduced by 52% in the axotomized cells when compared with control cells. Two current components were reduced after ligation, I(A) by 60%, I(K) by over 65%, compared with control cells. I(D) appeared unaffected after acute ligation. These results indicate a large reduction in overall K(+) current, resulting from reductions in I(K) and I(A), on large cutaneous afferent neurons after nerve ligation and have implications for excitability changes of injured primary afferent neurons.

Gradual Emergence of Song Selectivity in Sensorimotor Structures of the Male Zebra Finch Song System

Birdsong is a model system for understanding how motor and sensory information interact to coordinate behavior. Neurons in one potential site of sensorimotor integration, the forebrain nucleus HVc, have premotor activity during singing and auditory activity during playback of the bird's own song. It is not known whether the high degree of selectivity for learned features of song observed during playback arises in HVc or also in structures afferent to HVc. We recorded in anesthetized adult zebra finches from two structures afferent to HVc: either the nucleus interfacialis (NIf) or the L1 subdivision of the field L complex, and simultaneously from a second electrode in HVc. Correlations in the bursting pattern of ongoing activity of HVc and NIf recordings were observed; these helped to localize the first electrode to NIf recording sites. Most NIf neurons exhibited song-selective responses, but as a population, they were less selective than were HVc neurons. Most L1 neurons were not song-selective. NIf neurons have also been reported to have premotor activity during singing; thus, NIf is another potential site of auditory-motor interactions in the song system. Evidence gathered to date suggests that those brain areas in the passerine forebrain that are recruited during song production also display the most selective learned auditory responses.

Increase of galanin mRNA in lumbar dorsal root ganglion neurons of adult rats after partial sciatic nerve ligation

Partial sciatic nerve ligation (PSNL) is a widely used model for the study of neuropathic pain. However, there is little information on neuropeptide expression in primary sensory neurons after PSNL. We examined galanin (GAL) mRNA expression in L4 and L5 dorsal root ganglion (DRG) neurons of adult rats after PSNL. We found that 4 and 14 days after PSNL the percentages of GAL mRNA positive neurons were significantly increased in the ipsilateral DRG compared to the contralateral side. Using combined retrograde fluorescent dye tracing and in situ hybridization, we found that 47% of the injured neurons and 10% of the spared neurons were GAL mRNA positive. Since only 2–3% of neurons in the contralateral uninjured DRG were GAL mRNA positive, PSNL induced up-regulation of GAL mRNA in both injured and spared DRG neurons.

Change in mRNAs for neuropeptides and the GABA A receptor in dorsal root ganglion neurons in a rat experimental neuropathic pain model

We examined two possible mechanisms of neuropathic pain: contribution of adjacent intact nerves and decrease in presynaptic inhibition at the central terminal of the injured primary afferent. To this end, we examined the effects of unilateral L5 spinal nerve ligation, which causes mechanical allodynia and heat hyperalgesia in the ipsilateral hind paw, on gene expression in L4 and L5 dorsal root ganglion (DRG) neurons using in situ hybridization (ISH). Specifically, we examined changes in the expression of messenger RNAs (mRNAs) for neuropeptides which have been reported to be up- or down-regulated in the axotomized DRG neurons and for γ-aminobutyric acid (GABA) A receptor (GABA A-R) subunits which contribute to presynaptic inhibition at the primary afferent terminals. Seven days following ligation, ISH demonstrated an increase in signal intensity for calcitonin gene-related peptide (CGRP) mRNA in the subpopulation of small- to medium-sized L4 DRG neurons ipsilateral to the ligation which were not directly injured as compared to the contralateral side, although the overall percentages and the size distribution of positively labelled neurons for CGRP mRNA were not different between the bilateral L4 DRGs. This suggests that the L4 DRG neurons which express CGRP mRNA constitutively up-regulated the gene expression and the functional importance of these neurons has increased following L5 spinal nerve ligation. However, the mRNAs for other neuropeptides such as preprotachykinin (PPT), vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), and galanin (GAL), were not different between the bilateral L4 DRGs. The mRNA for the GABA A-R γ2 subunit was significantly down-regulated in the medium- to large-sized L5 DRG neurons ipsilateral to the ligation as compared to the contralateral side. GABA A-R α2 subunit mRNA also decreased in the ipsilateral L5 DRG neurons but did not reach statistical significance. There was no difference in mRNAs between the bilateral L4 DRGs. These data suggest that the presynaptic disinhibition of the ipsilateral L5 primary afferent terminals may be explained at least partly by the down-regulation of GABA A-R following L5 spinal nerve ligation. Thus, both the up-regulation of CGRP in adjacent intact nerves and the decrease in presynaptic inhibition at the central terminal of the injured primary afferent could cause the hyper-excitability of dorsal horn neurons and contribute to the molecular mechanisms of this neuropathic pain model.

Effects of ethanol on rat somatosensory cortical neurons

In this study, we characterized the local effects of ethanol (EtOH) on postsynaptic potentials (PSPs) and membrane properties of layer II–III (L2–3) and layer V (L5) somatosensory cortical neurons. Intracellular recordings were done using the in vitro slice preparation of rat somatosensory cortex. Our results show that EtOH exerts local effects on cortical cell membrane at physiologically relevant concentrations. A predominant effect of EtOH was to reduce excitability of L2–3 and L5 neurons by increasing the rheobase, decreasing input resistance and repetitive firing, reducing PSPs amplitude and the probability of evoking action potentials. Early (6 ms) and late (18 ms) PSP components were affected differentially by EtOH, the late components being more suppressed. Overall, EtOH-mediated suppression of PSPs was stronger in L5 neurons. Cortical neurons were divided into three subtypes: regular spiking adapting (RS-A), regular spiking non-adapting (RS-NA) and bursting (D-IB) neurons. PSPs evoked in RS-A neurons were more sensitive to EtOH suppressant effects. EtOH effects on input resistance were distributed differentially among the three groups of neurons. These results support the notion that EtOH disrupts higher processing of somatosensory information via a differential alteration of cortical neuron's membrane properties and synaptic transmission.

Inhibition of a hyperpolarization-activated current by clonidine in rat dorsal root ganglion neurons.

Whole cell voltage- and current-clamp recordings were carried out to investigate the effects of clonidine, an alpha 2-adrenoceptor agonist, in L4 and L5 dorsal root ganglion (DRG) neurons of the rat. In voltage-clamp mode, application of 20 microM clonidine reversibly reduced the inward current evoked by hyperpolarizing voltage steps. The "clonidine-sensitive current" was obtained by subtracting the current during clonidine application from the control current, and its properties were as follows. 1) It was a slowly activating inward current evoked by hyperpolarization. 2) The reversal potential in the standard extracellular solution ([K+]o = 5 mM, [Na+]o = 151 mM) was -38.3 mV, and reduction of [Na+]o shifted it to a more negative potential, whereas an increase of [K+]o shifted it to a more positive potential, indicating that the current was carried by Na+ and K+ (PNa/PK = 0.22). 3) The relationship between the chord conductance underlying the clonidine-sensitive current and voltage could be fitted by a Boltzmann equation. These results indicate that the clonidine-sensitive current corresponds to a hyperpolarization-activated current (Ih), i.e., clonidine inhibits Ih in rat DRG neurons. DRG neurons were classified as small (15.9-32.9 microns diam), medium-sized (33-42.9 microns), and large (43-63.6 microns), and 7 of 19, 24 of 25, and 22 of 22 of these types exhibited Ih with mean +/- SE clonidine-induced inhibition values of 36.1 +/- 3.5% (n = 7), 43.1 +/- 3.7% (n = 24), and 35.1 +/- 2.7% (n = 22), respectively. Clonidine application to L4 and L5 DRG neurons excised from rats the sciatic nerves of which had been transected 14-35 days previously (transected DRG neurons) also reduced Ih. In current-clamp mode, 9 of 13 intact and 4 of 6 transected medium-sized DRG neurons that exhibited Ih responded to clonidine with hyperpolarization (> 2 mV). Some medium-sized DRG neurons exhibited repetitive action potentials in response to a depolarizing current pulse, and clonidine reduced the firing discharge frequencies in 8 of 11 intact and 3 of 4 transected neurons tested. Injection of a hyperpolarizing current pulse produced time-dependent rectification in DRG neurons that exhibited Ih, and clonidine blocked this rectification in all intact and transected neurons tested. These results suggest that inhibition of Ih due to alpha 2-adrenoceptor activation contributes to modulation of DRG neuronal activity in rats. On the basis of our findings, we discuss the possible mechanisms whereby sympathetically released norepinephrine modulates the abnormal activity of DRG neuronal cell bodies after nerve injury.

Increase of preprotachykinin mRNA and substance P immunoreactivity in spared dorsal root ganglion neurons following partial sciatic nerve injury.

Complete sciatic nerve injury reduces substance P (SP) expression in primary sensory neurons of the L4 and L5 dorsal root ganglia (DRG), due to loss of target-derived nerve growth factor (NGF). Partial nerve injury spares a proportion of DRG neurons, whose axons lie in the partially degenerating nerve, and are exposed to elevated NGF levels from Schwann and other endoneurial cells involved in Wallerian degeneration. To test the hypothesis that SP is elevated in spared DRG neurons following partial nerve injury, we compared the effects of complete sciatic nerve transection (CSNT) with those of two types of partial injury, partial sciatic nerve transection (PSNT) and chronic constriction injury (CCI). As expected, a CSNT profoundly decreased SP expression at 4 and 14 days postinjury, but after PSNT and CCI the levels of preprotachykinin (PPT) mRNA, assessed by in situ hybridization, and the SP immunoreactivity (SP-IR) of the L4 and L5 DRGs did not decrease, nor did dorsal horn SP-IR decrease. Using retrograde labelling with fluorogold to identify spared DRG neurons, we found that the proportion of these neurons expressing SP-IR 14 days after injury was much higher than in neurons of normal DRGs. Further, the highest levels of SP-IR in individual neurons were detected in ipsilateral L4 and L5 DRG neurons after PSNT and CCI. We conclude that partial sciatic nerve injury elevates SP levels in spared DRG neurons. This phenomenon might be involved in the development of neuropathic pain, which commonly follows partial nerve injury.

Correlation of nicotinic receptor-like mRNA expression with excitatory input into the behaviorally important L1 and L3 auditory interneurons of the cricket,Acheta domesticus

Nonradioactive in situ hybridization was used on sections of the prothoracic ganglion of the female cricket Acheta domesticus, to evaluate expression of an mRNA targeted by alkaline phosphatase-linked oligonucleotide probes based on the sequence of the α-L1 subunit of the locust nicotinic acetyl choline receptor. Somata for the L1 and L3 auditory interneurons (previously shown to be behaviorally important in the phonotactic response of females to the males' calls) in the prothoracic ganglion were stained with experimental probes for two different nucleotide sequences within this gene. Controls (heterologous probe targeting other mRNA, sense control probe, control probe for specific nucleotide sequences, competition control, and absence of probe) produced no evidence of hybridization in L1 or L3 neuron somata. The two experimental probes showed hybridization patterns in the L1 and L3 neurons that correlated well the neuron's level of excitation and with the female's phonotactic behavior. The hybridization signal is high in the L1 neuron somata of females at ages (4–28 days old) when phonotactic and L1 thresholds are low. Females (4–12 days old) that typically were selective for the syllable period of the male's calling song and showed higher levels of excitation in their L3 neurons demonstrated higher hybridization in the somata of these neurons. J. Exp. Zool. 281:109–123, 1998. © 1998 Wiley-Liss, Inc.

Morphological and morphometrical changes in dorsal root ganglion neurons innervating the regenerated lizard tail

The variations occurring in neurons from dorsal root ganglia that provide innervation to the regenerated tail of the lizard (vicarious ganglia) are analysed. Vicarious ganglion neurons, when compared to control ganglion neurons (i.e. ganglia from the same animal that were not involved in the reinnervation process), show a size increase of the soma (cell hypertrophy) which applies to all cell types and sub-types. No statistically significant differences in the relative percentage of neurofilament-poor (type D) and neurofilament-rich (type L) neurons were found between vicarious dorsal root ganglia compared to controls in all animals. On the contrary, within L neuron sub-types, a statistically significant increase in sub-type L2 (very rich in neurofilaments), and the appearance of sub-type L3 neuron which is not detectable in controls, were demonstrated in vicarious dorsal root ganglia. In spite of these variations in size and percentage distribution, no structural and ultrastructural differences of the various cell types and sub-types are detectable, except for the appearance of the sub-type L3 neurons. However, this neuron sub-type might not be considered specific of hypertrophy since the same morphological features have been observed, in normal conditions, in lizard dorsal root ganglia from cervical and lumbar spinal levels that provide innervation to limb plexuses.

Reduced nuclear factor kappaB (p65) expression in rat primary sensory neurons after peripheral nerve injury.

The activated form (p65) of nuclear factor kappa B (NF-kappa B) was found to be expressed in a sub-population (32%) of mixed diameter rat sensory neurons in L4 and L5 dorsal root ganglia, indicating that this transcription factor is involved in intracellular signalling in sensory neurons under physiological conditions. Four hours after crushing the sciatic nerve, ipsilateral p65 staining was abolished in a subgroup (60-70%) of these neurons. The contralateral side was unaffected by the injury and loss of NF-kappa B activity was not observed following sham surgery. Within 24 h of sciatic injury, ipsilateral p65 staining had recovered to control levels. We suggest that the decline in p65 after nerve injury was due to failure of retrograde axonal transport of trophic factor(s). Since neurotrophins and cytokines promote the survival of non-neuronal cells by activation of NF-kappa B, we believe that p65 may be critical for the resistance to apoptosis shown by adult sensory neurons.

Emerging auditory response interactions to harmonic complexes in field L of the zebra finch

Auditory responses of single units to harmonic complexes were studied in field L of the adult male zebra finch (Poephila guttata) to understand the origin of song selectivity in HVc. One of the attributes in this song selectivity is a large increase in response interactions when tones are presented simultaneously. Single units in the subdivisions of field L-L1, L2, and L3, were examined to see where these response interactions to harmonic complexes begin to develop along the auditory pathway towards HVc. Results showed relatively simple stereotyped responses in L2 and widely varying ones in L1 and L3. The responses of some neurons in L1 and L3 to signals composed of the simultaneous sum of two harmonic complexes significantly differed from the linear sum of their responses to individual harmonic complexes whereas the neurons of L2 showed relatively weak response interactions. These neurons in L1 and L3 exhibited strong response interactions to harmonic complexes comparable to those of song-selective neurons in HVc. Thus, L1 and L3 are suggested to have emerging selective response properties and to provide auditory inputs relevant to the song selective HVc neurons.

Sensory innervation to the anterior portion of lumbar intervertebral disc.

The level of dorsal root ganglia that receives sensory afferent nerves from the anterior portion of the lower lumbar intervertebral disc was investigated in rats using a retrograde transport method. Sometimes patients with lower lumbar disc lesions complain of inguinal pain that does not correspond to the dermatome of the injured nerve roots. To investigate the origin of the pain, the authors studied the sensory innervation to the anterior portion of the lumbar intervertebral disc. The innervation to the posterior portion of the lumbar disc has been extensively investigated and has been reported to be segmental. However, little is known about the nerve supply to the anterior portion of the lumbar disc. The retrograde transport method was used in rats. As tracers, horseradish peroxidase and choleratoxin B subunit were used. Horseradish peroxidase crystals were placed on the anterior portion of the L5-L6 disc, and choleratoxin B subunit was injected into the L5-L6 disc. The bilateral dorsal root ganglia were histologically examined. Labeling of L1 and L2 dorsal root ganglia neurons was recognized. No neurons were labeled in dorsal root ganglia of other levels, including the segmentally corresponding L5. Using the retrograde transport method, the authors demonstrated that the anterior portion of the L5-L6 lumbar intervertebral disc was innervated from L1 or L2 spinal nerves in rats. These results appear to explain the reason why patients with lower lumbar disc lesions sometimes complain of inguinal pain corresponding to the L1-L2 dermatome.

Parallel pathways and convergence onto HVc and adjacent neostriatum of adult zebra finches (Taeniopygia guttata).

The structure and connectivity of the forebrain nucleus HVc, a site of sensorimotor integration in the song control system of oscine birds, were investigated in adult zebra finches. HVc in males comprises three cytoarchitectonic subdivisions: the commonly recognized central region with large and medium-sized darkly staining cells, a ventral caudomedial region with densely packed small and medium-sized cells, and a dorsolateral region with oblong cells and rows of cells. All three subdivisions project to area X and the robust nucleus of the archistriatum, with more complexity in the classes and distribution of cells than previously reported. In females, HVc is very small and has a cytoarchitecture distinct from that of the three male subdivisions. The structure of HVc in females treated with estradiol at 15 days of age is similar to male HVc. Tracer studies in males with fluorescent and biotinylated dextrans demonstrate non-topographic projections onto HVc that may carry auditory information, including type 1 and type 2 neurons in subdivisions L1 and L3 of the field L complex, a class of neurons in nucleus interface, nucleus uvaeformis, the caudal neostriatum ventral to HVc, and intrinsic HVc connections. These data are interpreted in terms of HVc's functional properties. Additionally, the neostriatum immediately ventral to HVc receives projections from field L, ventral hyperstriatum, and caudal neostriatum, and projects to a region surrounding RA and near to or into area X. The similarity of the connectivity of HVc and adjacent neostriatum suggests the possibility that they share a common origin.

Up-Regulation of Laminin B2 Gene Expression in Dorsal Root Ganglion Neurons and Nonneuronal Cells during Sciatic Nerve Regeneration

A recent study in our laboratory showed that B2 laminin gene is expressed by all L4 and L5 dorsal root ganglion (DRG) neurons as well as by satellite and Schwann cells. Because the laminin B2 subunit has a domain that supports neurite extension in culture, the present study was undertaken to test the hypothesis that laminin B2 gene expression would increase during sciatic nerve regeneration. In situ hybridization was used to examine B2 laminin gene expression in L4 and L5 DRGs 28 days after creating and bridging a 10-mm sciatic nerve gap with an impermeable silicone tube. Overall there was a nearly threefold increase in DRG B2 laminin chain mRNA at this timepoint, a time when axons are known to show vigorous regrowth. Both neurons and nonneuronal cells contributed to this increase. These data suggest that an up-regulation of B2 laminin gene expression by DRG neurons and nonneuronal cells may play a role in peripheral nerve regeneration.

Neuropeptide Y and galanin binding sites in rat and monkey lumbar dorsal root ganglia and spinal cord and effect of peripheral axotomy.

Using monoiodinated peptide YY (PYY) and galanin as radioligands, and neuropeptide Y (NPY) fragments, the distribution of NPY binding sites and its subtypes Y1 and Y2, and of galanin binding sites, was investigated in rat and monkey lumbar (L) 4 and L5 dorsal root ganglia (DRG) and spinal cord before and after a unilateral sciatic nerve cut, ligation or crush. Receptor autoradiography revealed that [125I]PYY bound to some DRG neurons and a few nerve fibres in normal rat DRG, and most of these neurons were small. NPY binding sites were observed in laminae I-IV and X of the rat dorsal horn and in the lateral spinal nucleus, with the highest density in laminae I-II. [125I]PYY binding was most strongly attenuated by NPY13-36, a Y2 agonist, and partially inhibited by [Leu31,Pro34]NPY, a Y1 agonist, in both rat DRG and the dorsal horn of the spinal cord. These findings suggest that Y2 receptors are the main NPY receptors in rat DRG and dorsal horn, but also that Y1 receptors exist. After sciatic nerve cut, PYY binding markedly increased in nerve fibres and neurons in DRG, especially in large neuron profiles, and in laminae III-IV of the dorsal horn, as well as in nerve fibres in dorsal roots and the sciatic nerve. Incubation with NPY13-36 completely abolished PYY binding, which was also reduced by [Leu31,Pro34] NPY. However, the increase in PYY binding seen in laminae I-IV of the ipsilateral dorsal horn after axotomy was not observed after coincubation with [Leu31,Pro34] NPY. NPY binding sites were seen in a few neurons in monkey DRG and in laminae I-II, X and IX of the monkey spinal cord. The intensity of PYY binding in laminae I-II of the dorsal horn was decreased after axotomy. Galanin receptor binding sites were not observed in rat DRG, but were observed in the superficial dorsal horn of the spinal cord, mainly in laminae I-II. Axotomy had no effect on galanin binding in rat DRG and dorsal horn. However, galanin receptor binding was observed in many neurons in monkey L4 and L5 DRG and in laminae I-IV and X of monkey L4 and L5 spinal cord, with the highest intensity in laminae I-II. No marked effect of axotomy was observed on the distribution and intensity of galanin binding in monkey DRG or spinal cord.(ABSTRACT TRUNCATED AT 400 WORDS)

Some sensory neurons express neuropeptide Y receptors: potential paracrine inhibition of primary afferent nociceptors following peripheral nerve injury

Neuropeptide Y (NPY) has been suggested to exert antinociceptive actions by inhibiting the release of neurotransmitters from trigeminal and dorsal root ganglia (DRG) neurons, but the site of direct NPY action in vivo and the NPY receptor subtype mediating these effects are unknown. 125I-peptide YY (PYY) was used to localize and characterize NPY receptor binding sites in trigeminal ganglia, DRG, and spinal cord of the rat, rabbit, and monkey. In the rat, rabbit, and monkey, 5-20% of trigeminal ganglia and DRG neurons express NPY binding sites. Unilateral cuff-induced neuropathy or transection of the rat sciatic nerve did not significantly alter the density or number of DRG neurons expressing NPY receptors. A unimodal size distribution for L4 and L5 DRG neurons expressing NPY binding sites in the rat was determined, with a mean cross-sectional area of 947 microns 2. In the spinal cord the highest concentration of NPY receptors is found in laminae I, II, V, X, and Onuf's nucleus. Pharmacological experiments using selective Y1 and Y2 receptor antagonists suggest that Y2 is the prominent NPY receptor subtype expressed in trigeminal ganglia neurons, DRG neurons, and spinal cord. Previous studies have demonstrated that a population of large-diameter, presumably myelinated primary afferents express NPY after peripheral nerve injury. NPY released from these injured large-diameter DRG neurons may act in a paracrine fashion to block the transmission of nociceptive information from the small- and medium-diameter DRG neurons that constitutively express NPY receptors. NPY receptors are therefore uniquely positioned to inhibit primary afferent nociceptors directly, especially after peripheral nerve injury.

Lox1, an Antennapedia-Class Homeobox Gene, Is Expressed during Leech Gangliogenesis in Both Transient and Stable Central Neurons

The leech homeobox gene Lox1 contains a homeodomain homologous to those of Sex combs reduced and Antennapedia of Drosophila, and to homeodomains of the vertebrate Hox groups 5 and 6. Lox1 expression was detected during late gangliogenesis in 15-20 pairs of central neurons repeated in most segments, suggesting that Lox1 may play a role in the differentiation of these cells. The monoclonal antibody Laz1-1 identified two pairs of Lox1-expressing neurons as the Bipolar cells and the L1 neurons. The L1 neurons were detected late in gangliogenesis and became stable neurons. The Bipolar cells appeared very early in gangliogenesis, extended processes in the longitudinal connective nerves, and then degenerated, suggesting that they play a role in establishing the longitudinal tracts of the central nerve cord.

Antennapedia-class homebox genes define diverse neuronal sets in the embryonic CNS of the leech.

Studies of the Antennapedia-class homeobox genes suggest that specific combinations of these transcription factors play a role in defining neuronal identities. We examined the expression of these genes in the leech Hirudo medicinalis, an organism well-suited for neurobiological research at the level of identified neurons. Leeches contain at least as many Antennapedia-class and related genes as insects do, despite the apparently lower complexity of the leech body plan. The CNS expression patterns of two Antennapedia-class leech homeobox genes (Lox genes) were examined in detail. Lox1 is expressed during early gangliogenesis in only one pair of transient neurons present in every segment (the Bipolar cells) and, at later stages of embryonic development, in 15-20 pairs of central neurons repeated in most segments. The monoclonal antibody Laz1-1 identified two pairs of Lox1-expressing neurons as the Bipolar cells and the L1 neurons. The Bipolar cells extended processes in the primordia of the longitudinal connective nerves and later degenerated. The L1 neurons were detected late in gangliogenesis and became stable neurons. Lox2 is expressed in an iterated set of neurons in the posterior two-thirds of the CNS. On the basis of cell body position and relative size, two pairs of Lox2-expressing cells were identified as the RPE-like neurons and the CV motor neurons. Other Lox genes are also expressed in segmentally repeated subpopulations of neurons. These neuronal subpopulations appear to be different from one another but partially overlapping. Different combinations of Lox genes that may be expressed in individual cells could in theory generate enough variability to specify all central neurons in a leech ganglion.

Intraspinal modulation of neuronal responses to uterine and cervix stimulation in rat L1 and L6 dorsal horn

This two-part study examined intraspinal processing of input from the uterus, cervix and skin within the caudal spinal cord of virgin rats in estrus. The first part examined the effects of removing either pelvic or hypogastric nerve input (by appropriate dorsal rhizotomies) on neuronal responses to stimulation of uterus, cervix and skin in either the L1 or L6 dorsal horn of decerebrate, spinalized unanesthetized rats. Bilateral section of the T13-L2 roots (through which most hypogastric afferents travel) eliminated almost all responses to uterine distension in L1 and L6. Such rhizotomies also decreased the inhibitory effects of cervix input on neurons in L6. Bilateral section of the L6-S2 roots (through which most pelvic afferents travel) had no effect on responses to uterus, but decreased the inhibitory effects of cervix input on neurons in L1. Sections of both sets of roots eliminated responses to cervix stimulation in L6. The second part examined neuronal response in T13/L1 of urethane-anesthetized rats before and after a T10 spinal transection. Transection increased the probability of observing neurons excited by uterine stimulation. Transection also increased excitatory responses and decreased inhibitory responses to cervix stimulation. These results confirm previous findings that input from uterus to the spinal cord is mainly by way of the hypogastric nerve, while that from the cervix is by way of both the pelvic and hypogastric nerves. The results also demonstrate that descending influences in the estrous rat mainly increase the ability of cervix stimulation to inhibit spinal neuronal activity and reduce the effectiveness of uterine stimulation. Intersegmental effects between L1 and L6 as well as effects from distant roots within each segment on the processing of input from the cervix are similarly generally inhibitory. The challenge now is to understand how these interacting, mainly inhibitory effects sculpt the actions of neurons for various aspects of reproduction and other functions.