Initial trajectories of sensory axons toward laminar targets in the developing mouse spinal cord

Quantitative ultrasonic tissue characterization using backscattered high-frequency intravascular ultrasound could provide a basis for the objective identification of lesions in vivo. Representation of local measurements of quantitative ultrasonic parameters in a conventional image format should facilitate their interpretation and thus increase their clinical utility. Toward this goal, the apparent integrated backscatter, the slope of attenuation (25-56 MHz) and the value of the attenuation on the linear fit at 37.5 MHz were measured using the backscattered radio frequency signals from in vitro human aortae. Local estimations of these ultrasonic parameters from both normal and atherosclerotic aortic segments were displayed in a B-scan format. The morphological features of these parametric images corresponded well to features of histological images of the same regions. The attenuation from 25-56 MHz of seven segments of the medial layer (both with and without overlying atheroma) were measured using the multinarrow-band backscatter method. The average attenuation in the media at 24 degrees C +/- 3 degrees C was 45 +/- 16 dB/cm at 25 MHz and 102 +/- 13 dB/cm at 50 MHz. This work represents progress toward the development of quantitative imaging methods for intravascular applications.     

Regeneration of ascending spinal axons in goldfish

Regeneration of descending spinal cord tracts occur spontaneously in adult goldfish. Very little information is available regarding the fate of ascending fibers. Using Dextran amines as a tracer, we studied the normal and regenerated ascending axonal projection patterns in adult goldfish brain nuclei. Present study includes spinal projections to torus semicircularis, hypothalamus, thalamus and the telencephalon. Regenerated fibers had finer caliber axons and the terminal axonal arbors covered a larger area than the corresponding normal ones.     

Immunohistochemical localization of enkephalin in peripheral sensory axons in the rat

Impaired energy metabolism plays an important role in neuronal cell death after brain ischemia, and apoptosis has been implicated in cell death induced by metabolic impairment. In the present study, metabolic impairment was induced by 3-nitropropionic acid (3-NP), an irreversible inhibitor of succinate dehydrogenase. In order to clarify the involvement of poly(ADP-ribosyl)ation and apoptotic pathway in 3-NP induced cell death, we examined poly(ADP-ribosyl)ation and the apoptosis related gene protein expression after systemic administration of 3-NP by immunohistochemistry. Poly(ADP-ribosyl)ation was evidently detected in the striatal lesion but not in any other region. Immunoreactive ratio of Bcl-2 to Bax significantly increased both in the striatum and cortex. The data suggest that striatal cell death involves poly(ADP-ribosyl)ation and also apoptotic pathway in part following administration of 3-NP.     

Gene-expression screens in vertebrate embryos: more than meets the eye

The health control of the travellers and medical assistance for them and their families is one of the oldest and most fundamental missions of the "Office des migrations internationales" (OMI) as defined by the reglementary treaties of November 1945 and June 1946. More recent reglementations have broadened the competence of the OMI to include certain categories of foreigners who had been exempt from all controls. Finally, the ministerial order of November the 7th 1994 specifies means of the health screening. Medical examinations are carried in various French and foreign institutions. It nowadays has become a consultation aiming at prevention and orientation. The medical examination has three principal objectives: the detection of little known abnormalities; the contact between the medical service of the OMI and regional services in charge of the first check-up; the health education of the examined persons. The clinical and paraclinical results are communicated to the patient who is also informed of the most serious health questions, as well as ways of gaining access to medical care in France; this is done by taking into account all medical parameters and health conditions prevailing in the patient's native country. All detected abnormalities are brought to the attention of the head physician of the OMI who in turn informs the medical inspector of the "Direction départementale des affires sanitaires et sociales" (DDASS) in charge of making sure that the migrant benefits from health and social assistance and receives medical treatment. All pathological results are given to the examined persons in form of a written and confidential report, enabling them to visit a doctor of their choice. A network has thus been built up throughout the various departments involved in the first medical examination and the DDASS has made available for the OMI medical staff listings of public institutions likely to welcome the migrants.     

Selective disruption of neuregulin-1 function in vertebrate embryos using ribozyme-tRNA transgenes

The products of the neuregulin-1 gene constitute a set of polypeptide growth factors whose signalling through the ErbB receptors is essential to the growth and differentiation of many cell types in culture. Although studies with neuregulin-1 mutant mice have demonstrated that these growth factors are also essential regulators of cellular differentiation in vivo, the mid-embryonic death of these mutants precludes an analysis of hypothesized neuregulin-1 roles in later aspects of development. To circumvent this early lethality, we have pursued a ribozyme-based strategy for the perturbation of neuregulin-1 function in developing chick embryos. Early administration of a retrovirus carrying neuregulin-1 hammerhead-type ribozymes to blastoderm-stage embryos leads to an embryonic lethal phenotype that results from the failure of ventricular trabeculation in the developing heart, a faithful phenocopy of the mouse neuregulin-1 mutations. Later, more localized delivery of the ribozyme to the developing retina inhibits both the differentiation of retinal ganglion cell neurons and the proliferation of the neuroepithelial cells from which they derive. These results suggest that neuregulin-1 promotes both muscle cell differentiation in the heart and neuronal differentiation in the central nervous system. In addition, they demonstrate the utility of hammerhead ribozymes as a simple, effective and easily adaptable method of conditional gene inactivation in vertebrates.     

Fixation of spinal reflex alterations in spinal rats by sensory nerve stimulation.

Two preparations in which sensory nerve stimulation was used to obtain peripherally induced spinal fixation in spinal rats are described. In the first preparation, proportionally greater amounts of persisting poststimulation flexor muscle contraction, as measured by a force displacement transducer, were produced as stimulation time was increased from 10 min to 40 min. In the second preparation, sensory nerve stimulation was delivered, and evoked whole-nerve responses were recorded from a flexor motor nerve. Results indicated that 30 min or more of sensory nerve stimulation produced increases in response amplitude and area that persisted for at least 30 min after stimulation. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Kainate-receptor-mediated sensory synaptic transmission in mammalian spinal cord

Glutamate, the major excitatory neurotransmitter in the central nervous system, activates three different receptors that directly gate ion channels, namely receptors for AMPA (alpha-amino-3-hydroxy-5-methyl isoxozole propionic acid), NMDA (N-methyl-D-aspartate), and kainate, a structural analogue of glutamate. The contribution of AMPA and NMDA receptors to synaptic transmission and plasticity is well established. Recent work on the physiological function of kainate receptors has focused on the hippocampus, where repetitive activation of the mossy-fibre pathway generates a slow, kainate-receptor-mediated excitatory postsynaptic current (EPSC). Here we show that high-intensity single-shock stimulation (of duration 200 microseconds) of primary afferent sensory fibres produces a fast, kainate-receptor-mediated EPSC in the superficial dorsal horn of the spinal cord. Activation of low-threshold afferent fibres generates typical AMPA-receptor-mediated EPSCs only, indicating that kainate receptors may be restricted to synapses formed by high-threshold nociceptive (pain-sensing) and thermoreceptive primary afferent fibres. Consistent with this possibility, kainate-receptor-mediated EPSCs are blocked by the analgesic mu-opiate-receptor agonist Damgo and spinal blockade of both kainate and AMPA receptors produces antinociception. Thus, spinal kainate receptors contribute to transmission of somatosensory inputs from the periphery to the brain.     

Guidance of circumferentially growing axons by netrin-dependent and -independent floor plate chemotropism in the vertebrate brain

BACKGROUND: A strong positive correlation exists between the breast cancer tissue content of either urokinase-plasminogen activator (uPA) or plasminogen activator, inhibitor type I (PAI-1), quantified in the tissue extracts by immunoassays, and the survival of patients with breast cancer. Furthermore, several studies assign to the urokinase-type plasminogen activator receptor (uPAR) a pivotal role in triggering the proteolytic activity of the urokinase pathway involved in tumor stroma degradation, tumor spread and metastasis. However, the pattern of distribution of uPAR in normal and cancerous human tissue and the pattern of coexpression of activators and inhibitors that occurs in breast cancer tissues is not completely known. METHODS: The immunohistochemical localization of uPAR, uPA, tPA) and PAI-1 was evaluated by using the avidin-biotin immunoperoxidase technique and affinity-purified monoclonal antibodies from American Diagnostica Inc. Studies were performed in formalin fixed, paraffin-embedded tissue prepared from 23 surgically excised non-neoplastic breast tissues and 18 ductal breast carcinomas. RESULTS: While the expression of uPAR protein represents a constant feature of invasive ductal breast cancer, it was also observed in most of the breast tissue samples, including the normal breast tissues. The staining for uPAR was mainly localized on normal or tumoral epithelial cells, even if the co-expression of uPAR in stromal cells was frequently observed in adjacent slides. A semiquantitative analysis of immunohistochemical results showed that uPAR and PAI-1 were overexpressed in invasive breast cancer in comparison with normal and benign breast tissues. In addition, uPA was higher in both invasive breast carcinomas and benign breast lesions with respect to normal breast tissues. CONCLUSIONS: We showed that overexpression of uPAR, uPA, and its main inhibitor, PAI-1, is a constant feature of invasive ductal breast carcinomas. However, the expression of the above fibrinolytic reactants is not specific for breast cancer since positive staining for these molecules was frequently observed in benign breast lesions as well as in normal breast tissues. The combined increased expression of uPA and its cellular receptor, uPAR on the surface of tumor epithelial cells may account for the activation of the proteolytic system which occurs in breast cancer.     

Postnatal growth of corticospinal axons in the spinal cord of developing mice

The corticospinal tract (CST) plays an important role in the control of voluntary movements. Although the development of the CST has been studied extensively in other species, limited information is available on its development in mice. In the present study, the growth of corticospinal axons was characterized in developing mice using Phaseolus vulgaris leucoagglutinin (PHA-L). Our results indicate that the leading CST axons reach the 8th cervical segment at postnatal day (PD) 2, the 7th thoracic segment at PD4, the 13th thoracic segment at PD7, and the 5th lumbar segment at PD9. The arrival of corticospinal axons at the distal lumbar cord at PD9 was further confirmed by retrograde tracing using fast blue (FB). A waiting period of 2-3 days exists after the leading CST axons pass a particular segment before sending collaterals into the gray matter of that segment. The CST continues to increase in size in lower thoracic and lumbar areas up to PD14 when its adult appearance is achieved. In this study, the date of animal's sacrifice was used as the specific postnatal date to demonstrate the growth of the CST. This definition gives a more reliable indication of the exact location of the CST at a specific developmental time point since the CST continues to grow after tracer injections and since the dye is transported much faster than axonal growth. We suggest that these findings can be used as a template for studies on both normal and transgenic mice where some developmental significance is given to the CST.     

Organization of primary afferent axons in the trigeminal sensory root and tract of the rat

A combination of immunocytochemical and electron microscopic methods were employed to assess the organization of the trigeminal (V) spinal tract in adult rats. Immunostaining was employed at the light microscopic level to selectively label large myelinated (by using antibodies against neurofilament protein) and small unmyelinated (by using antibodies against calcitonin gene-related peptide) primary afferents. In addition, the plant lectin Bandeiraea simplicifolia-I was employed to histochemically label small unmyelinated primary afferents. Results from these experiments indicated that larger myelinated axons were distributed throughout the cross-sectional extent of the V spinal tract (TrV), whereas smaller fibers were most numerous just below the pial surface. These results were confirmed with quantitative electron microscopy which demonstrated that the central portion of the V sensory root and TrV were composed primarily of larger myelinated fibers, whereas the periphery of the root and the portion of TrV just below the pial surface contained a higher percentage of smaller myelinated and unmyelinated axons. When considered together with results regarding the birthdates of neurochemically defined classes of V ganglion cells (White et al. [1994] J. Comp. Neurol. 350:397-411), these results suggest that TrV is laid down in a chronotopic fashion with the first axons forming its deeper portion and later arriving axons being added more superficially.     

Microtubule disorientation and axonal swelling in unmyelinated sensory axons during vincristine-induced painful neuropathy in rat

Neuropathic pain accompanies peripheral nerve injury following a variety of insults including metabolic disorders, traumatic injury, and exposure to neurotoxins such as vincristine and taxol. Vincristine, a microtubule depolymerizing drug, produces a peripheral neuropathy in humans that is accompanied by painful paresthesias and dysesthesias (Sandler et al., [1969] Neurology 19:367-374; Holland et al. [1973] Cancer Res. 33:1258-1264). The recent development of an animal model of vincristine-induced neuropathy provides an opportunity to investigate mechanisms underlying this form of neuropathic pain. Systemic vincristine (100 microg/kg) produces hyperalgesia to mechanical stimuli during the second week of administration, which persists for more than a week (Aley et al. [1996] Neuroscience 73:259-265). To test the hypothesis that changes in microtubule structure in nociceptive sensory neurons accompany vincristine-induced hyperalgesia, we analyzed unmyelinated axons in saphenous nerves of vincristine-treated rats. This study constitutes the first quantitative ultrastructural analysis of the cytoskeleton of unmyelinated axons in peripheral nerve during neuropathic hyperalgesia. There was no evidence of unmyelinated fiber loss or a decrease in the number of microtubules per axons. There was, however, a significant decrease in microtubule density in unmyelinated axons from vincristine-treated rats. This decrease in microtubule density was due to a significant increase in the cross-sectional area of unmyelinated axons, suggesting swelling of axons. In addition, vincristine-treated axons had significantly fewer microtubules cut in cross-section and significantly more tangentially oriented microtubules per axon compared to controls. These results suggest that vincristine causes disorganization of the axonal microtubule cytoskeleton, as well as an increase in the caliber of unmyelinated sensory axons.     

Sensory axons are guided by local cues in the developing dorsal spinal cord

During development, different classes of sensory neurons establish distinctive central projections within the spinal cord. Muscle spindle afferents (Ia fibers) grow ventrally through the dorsal horn to the ventral cord, whereas cutaneous sensory collaterals remain confined to the dorsal horn. We have studied the nature of the cues used by Ia fibers in establishing their characteristic projections within the dorsal horn. An organotypic culture preparation of embryonic chicken spinal cord and sensory ganglia was used to test the influence of ventral spinal cord and local cues within the dorsal spinal cord on the growing Ia afferents. When the ventral half of the spinal cord was replaced with an inverted duplicate dorsal half, Ia fibers entering through the dorsal columns still grew ventrally within the host dorsal horn. After the fibers entered the duplicate dorsal half, they continued growing in the same direction. With respect to the duplicate dorsal tissue, this was in an opposite, ventral-to-dorsal, direction. In both cases, however, Ia collaterals remained confined to the medial dorsal laminae. Restriction to these laminae was maintained even when the fibers had to change their direction of growth to stay within them. These results show that cues from the ventral cord are not required for the development of correct Ia projections within the dorsal horn. Local, rather than long-range directional, cues appear to determine the pattern of these projections. When the ventral half of the spinal cord was left intact but sensory axons were forced to enter the dorsal gray matter growing rostrally or caudally, their collateral axons grew in random directions, further showing the absence of directional cues even when the ventral cord was present. Taken together, these observations suggest that Ia fibers are guided by local positional cues that keep them confined to the medial gray matter within the dorsal horn, but their direction of growth is determined primarily by their orientation and position as they enter the dorsal gray matter.     

Effects of sex steroids on sensory and motor spinal mechanisms

The male copulatory pattern uses muscles in the penis for erection and penile insertion, the lower trunk for pelvic thrusting, and the sex accessory organs for seminal emission. Organization of the nuclei controlling penile muscles is achieved through cell growth, dendritic arborization, and synaptogenesis, actions dependent on androgen but not estrogen. Testosterone (T) and dihydrotestosterone (DHT) but not estradiol (E2), stimulate pelvic thrusting vigor by synchronizing discharge of motoneurons innervating pelvic muscles. Pelvic thrusting rhythmicity, regulated by spinal interneurons, is produced in female rabbits by E2 or T but not by DHT. Reflex contraction of the seminal vesicles, due to penile insertion, is facilitated by androgen presumably by its effect on preganglionic neurons of the hypogastric nerve, located in the dorsal commissural nucleus.     

Complex effects of general anesthesia on sensory processing in the spinal dorsal horn

A chronic animal preparation allowed us to compare activity of the same single, spinal dorsal horn neurons in the physiologically intact, awake, drug-free state and in the anesthetized state. The inhalation anesthetic enflurane produced profound, and at times, opposite effects on spinal dorsal horn neuron responses to non-noxious and noxious receptive field stimulation. Some effects would not have been predicted, based upon current understanding of anesthetics.     

Fixation of spinal reflexes in rats by central and peripheral sensory input.

Used 2 methods in 5 experiments (55 male hooded Long-Evans rats) to demonstrate retention of postural asymmetries after spinal cord section. In the 1st preparation, postural asymmetries of the hindlimbs were induced by placing electrolytic lesions in the anterior cerebellum. Asymmetry was found to consistently outlast a spinal cord section if 45 min were allowed between brain lesion and cord section. A certain percentage of Ss allowed 35 or 40 min also demonstrated the retention. In the 2nd preparation, postural asymmetries induced by 45 min of direct hindlimb stimulation were also retained after spinal section. Rhizotomy prior to stimulation resulted in a lack of appreciable asymmetry on termination of the stimulation. Retention of a hindlimb-stimulation-induced asymmetry was observed in Ss that underwent a spinal section before stimulation. Results demonstrate that the "spinal fixation" phenomenon can be obtained by induction of postural alterations from central (cerebellar lesion) and peripheral (hindlimb stimulation) sources. Results obtained from spinal Ss indicate that retention of peripherally induced asymmetry is not crucially dependent on higher brain center activity but rather seems to be more dependent on long-term alterations that occur directly in the spinal reflex system. (15 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

p75-deficient sensory axons are immunoreactive for the glycoprotein L1 in mice overexpressing nerve growth factor

Nerve growth factor (NGF) regulates the expression of the glycoprotein L1 among neural cell populations. The purpose of this investigation was to determine whether NGF equally affects the immunolocalization of L1 on both sympathetic and sensory axons, and whether the functional expression of the p75 neurotrophin receptor (p75NTR) is required for the immunodetection of this glycoprotein on peripheral axons. Two lines of transgenic mice overexpressing NGF among glial cells were used in this study: (1) one line of mice possessing two normal alleles for p75NTR, and (2) another line of mice possessing two mutated alleles for p75NTR. In both types of animals, sensory axons stained immunohistochemically for calcitonin gene-related peptide and sympathetic axons stained immunohistochemically for tyrosine hydroxylase invaded the deep white matter portions of the cerebellum (a central structure containing high levels of transgene expression and synthesis); the cerebella of wild type (C57Bl/6) and p75NTR-deficient mice lacked these sensory and sympathetic fibers. Both lines of transgenic animals also possessed a dense plexus of L1-immunoreactive axons in their cerebella; the spatial distribution of these L1-immunostained axons paralleled that seen for the sensory and sympathetic axons. A unilateral removal of the superior cervical ganglion in both lines of transgenic animals caused a complete reduction of tyrosine hydroxylase-immunopositive axons in the cerebellum but did not affect the density of L1-immunopositive axons. From these in vivo data, we conclude that collateral branches of sensory axons which invade a NGF-rich target area display L1 immunoreactivity, and that such immunodetection does not require the functional expression of p75NTR.     

Evidence for GAP-43 within descending spinal axons in the North American opossum, Didelphis virginiana

We have shown previously that GAP-43, a growth associated protein characteristically present in growing and regenerating axons, is relatively abundant in the spinal cord of adult opossums. In the present study, we combined the orthograde transport of the fluorescent marker Fluoro-Ruby with immunofluorescence for GAP-43 to determine if any of it is present within descending spinal axons. When Fluoro-Ruby was injected into the red nucleus and midbrain tegmentum, the medial pontine or medullary reticular formation, the medullary raphe or the lateral vestibular nucleus, axons were labeled in the expected areas of the spinal cord, but in most cases none showed evidence for GAP-43. In two of the four cases with rubral injections, however, a few labeled axons within the rubrospinal tract showed GAP-43 immunofluorescence, and in one case with an injection of the gigantocellular reticular nucleus and adjacent raphe, labeled axons within lamina IX immunostained for the protein. Since serotoninergic neurons are present within the gigantocellular reticular nucleus and adjacent raphe, and axons of the same phenotype are abundant within lamina IX, we asked whether serotoninergic axons contain GAP-43. When sections of the spinal cord were immunostained for both serotonin and GAP-43, many axons within lamina IX showed evidence for both substances. Such axons appeared to contact presumptive motoneurons. In cases with Fluoro-Ruby injections of the forelimb motor cortex, labeled axons were present within the pyramidal tract, and some of them showed evidence for GAP-43.     

Cholinergic contribution to excitation in a spinal locomotor central pattern generator in Xenopus embryos

1. We have investigated whether in Xenopus embryos, spinal interneurons of the central pattern generator (CPG) receive cholinergic or electrical excitatory input during swimming. The functions of cholinergic excitation during swimming were also investigated. 2. Intracellular recordings were made from rhythmically active presumed premotor interneurons in the dorsal third of the spinal cord. After locally blocking inhibitory potentials with 2 microM strychnine and 40 microM bicuculline, the reliability of spike firing and the amplitude of fast, on-cycle, excitatory postsynaptic potentials (EPSPs) underlying the single on-cycle spikes were measured during fictive swimming. 3. The nicotinic antagonists d-tubocurarine and dihydro-beta-erythroidine (DH beta E, both 10 microM) reversibly reduced the reliability of the spike firing during swimming and reduced the amplitude of the on-cycle EPSP by 16%. DH beta E also reduced the EPSP amplitude in spinalized embryos by 22%. These results indicate that interneurons receive rhythmic cholinergic excitation from a source within the spinal cord. 4. Combined applications of nicotinic and excitatory amino acid (EAA) antagonists or cadmium (Cd2+, 100-200 microM) resulted in complete block of the fast EPSP, suggesting that interneurons do not receive electrical excitation. 5. The nicotinic antagonists mecamylamine and d-tubocurarine (both 5 microM) reduced the duration of episodes of fictive swimming recorded from the ventral roots, in spinal embryos. When applied in the middle of a long episode, d-tubocurarine decreased the swimming frequency, ruling out an effect on the initiation pathway. The cholinesterase inhibitor eserine (10 microM) increased the duration of swimming episodes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Developmental plasticity of ascending spinal axons studies using the North American opossum, Didelphis virginiana

The polymerase chain reaction was used to characterize the trypanosomes infecting Glossina morsitans submorsitans and G. tachinoides in the game ranch of Nazinga, Burkina Faso, situated near an agropastoral zone. Dissection of 435 tsetse flies, and PCR analysis of 166 infected flies were conducted to assess the epidemiological situation. Trypanosomes of the Nannomonas subgenus were the most abundant in the two tsetse species (80.4% and 73.7% of identified infections in G. m. submorsitans and G. tachinoides respectively). T. vivax and T. brucei infection rates were comparable between the two tsetse species. Mature infection pattern identified by PCR differed from overall infections, mainly because T. simiae infections did not mature, whereas T. vivax represented the predominant taxon. Parasitological and PCR results showed some discrepancies; possibly some typical Duttonella strains could not be recognized by the sets of primers used. The technologies used in this work helped to determine the high trypanosomosis risk in this area.