Extracts of muscle from patients with amyotrophic lateral sclerosis enhance neurite outgrowth from ventral spinal cord explants of rat embryo

The effects of muscle extracts from normal controls and patients with amyotrophic lateral sclerosis (ALS) on neurite appearance in ventral spinal cord explants of 13-15-day-old Sprague-Dawley rats were investigated. Muscle extracts from normal controls and ALS muscle were significantly more effective in stimulating growth than control medium alone. There was no statistically significant difference between normal muscle extracts and ALS muscle extracts in growth of neurties. Our results may contribute to the hypothesis that ALS may be a disorder of motor neuron growth factors.     

Vasoactive intestinal peptide influences neurite outgrowth in cultured rat spinal cord neurons.

Vasoactive intestinal peptide (VIP) is a neuropeptide which has been shown to exhibit a wide range of neurotrophic effects both in vivo and in vitro. For the purpose of clarifying the effect of VIP on spinal cord neurons, we studied the effect of VIP on neurite outgrowth of fetal rat ventral and dorsal portions of spinal cord in cultures. VIP-treated ventral spinal cord cultures (VSCC), compared with control VSCC, had a significant neurite outgrowth at 10(-8), 10(-6), and 10(-4) M. The effect was considered to be concentration dependent. Morphological changes of the dorsal spinal cord cultures (DSCC) remained unchanged by VIP treatment. Because of their close sequence homology with VIP, PHI-27 (peptide, histidylisoleucine amide) and secretin were also examined with the same experimental conditions as was VIP. Both PHI-27 and secretin had neurite promoting effects in VSCC at 10(-8) and 10(-6) M, respectively. However, there were no neurite promoting effects in DSCC in both of them at any concentrations. VIP had the most potent effect on neurite outgrowth in VSCC, followed by PHI-27, and secretin in their effectiveness concentrations. Our data showing VIP, PHI-27 and secretin have neurotrophic action on VSCC and suggest that a potential therapeutic use of VIP and its related peptides in treating diseases that involve degeneration and death of spinal motor neurons, such as motor neuropathy and amyotrophic lateral sclerosis.     

促甲状腺释放激素类似物YM-14673对大鼠脊髓损伤后水肿的影响

目的研究促甲状腺释放激素（ＴＲＨ）类似物,ＹＭ－１４６７３大鼠脊髓损伤后水肿的影响。方法用改良Ａｌｌｅｎ氏法建立大鼠脊髓损伤模型,分设正常组、对照组和治疗组,治疗组在损伤后１５分钟注射ＹＭ－１４６７３,用称重法测量脊髓的水含量,公式：（湿重－干重）÷湿重×１００％。结果对照组示伤后２４小时脊髓水肿,治疗组显示在２４小时脊髓水肿减轻。结论早期应用ＴＲＨ类似物,ＹＭ-１４６７３可减轻脊髓损伤后的脊随水肿。     

Vasoactive intestinal peptide influences neurite outgrowth in cultured rat spinal cord neurons

Abstract

Vasoactive intestinal peptide (VIP) is a neuropeptide which has been shown to exhibit a wide range of neurotrophic effects both in vivo and in vitro . For the purpose of clarifying the effect of VIP on spinal cord neurons, we studied the effect of VIP on neurite outgrowth of fetal rat ventral and dorsal portions of spinal cord in cultures. VIP-treated ventral spinal cord cultures (VSCC), compared with control VSCC, had a significant neurite outgrowth at 10^-8, 10^-6, and 10^-4 M. The effect was considered to be concentration dependent. Morphological changes of the dorsal spinal cord cultures (DSCC) remained unchanged by VIP treatment. Because of their close sequence homology with VIP, PHI-27 (peptide, histidylisoleucine amide) and secretin were also examined with the same experimental conditions as was VIP. Both PHI-27 and secretin had neurite promoting effects in VSCC at 10^-8 and 10^-6 M, respectively. However, there were no neurite promoting effects in DSCC in both of them at any concentrations. VIP had the most potent effect on neurite outgrowth in VSCC, followed by PHI-27, and secretin in their effectiveness concentrations. Our data showing VIP, PHI-27 and secretin have neurotrophic action on VSCC and suggest that a potential therapeutic use of VIP and its related peptides in treating diseases that involve degeneration and death of spinal motor neurons, such as motor neuropathy and amyotrophic lateral sclerosis. [Neurol Res 2001; 23: 851-854]     

Tetrodotoxin enhances initial neurite outgrowth from fetal rat cerebral cortex cells in vitro

The effect of two different concentrations of the sodium channel blocker tetrodotoxin on neurite outgrowth from fetal rat cerebral cortex neurons was studied in vitro. A concentration of 10(-6) M tetrodotoxin enhanced after two days in vitro: the percentage of isolated cells and reaggregates which formed neurites, the number of neurites per cell soma, and neurite elongation and subsequent branching. The effects observed after treatment with 10(-7) M tetrodotoxin were on the whole intermediate between control and 10(-6) M tetrodotoxin.     

Effects of calcium ion on neurite outgrowth of rat spinal cord neurons in vitro: the role of non-neuronal cells in regulating neurite sprouting

The interactions of nerve cells with their environment and other cells are specific to different stages of cellular differentiation. Neurite outgrowth was measured from cultured spinal cord neurons under the influence of different Ca²⁺ concentrations. We used fluorodeoxyuridine (FuDr), an antimitotic agent which reduces significantly the proportion of non-neuronal cells in spinal cord cell cultures, to examine the effects of non-neuronal cells on neurite outgrowth. Spinal cord neurons responded to changes in their environment by means of two types of neurite outgrowth: sprouting and elongation. The concurrent presence of non-neuronal cells led to increased sprouting of neurites in certain ionic environments, thus lending support to the idea that non-neuronal cells release diffusible factors which influence sprouting and guide neurite outgrowth.     

Acidic and basic fibroblast growth factors enhance neurite outgrowth in cultured rat spinal cord neurons

Abstract

We have studied neurotrophic effects of acidic fibroblast growth factor (aFCF) and basic fibroblast growth factor (bFGF) on explanted ventral and dorsal spinal cord cultures from 13- and 14-day-old rat embryos. Cultures treated with aFCF and bFGF significantly enhanced neurite outgrowth with cultures of ventral spinal cord, but not with cultures of dorsal spinal cord. Our data suggest that aFCF and bFGF are potent neurotrophic factors on rat ventral spinal cord neurons in vitro. [Neurol Res 1995; 17: 70-72]     

Neurotrophic action of autopsied ventral spinal cord extracts in patients with amyotrophic lateral sclerosis on the ventral spinal cord of rat embryo

The effects of ventral spinal cord extracts from normal controls and patients with amyotrophic lateral sclerosis (ALS) on neurite appearance in ventral spinal cord explant of 13-day-old Sprague-Dawley rats were investigated. Ventral spinal cord extracts from normal controls and ALS were significantly more effective in stimulating growth than control medium only. There was no statistically significant difference between normal and ALS ventral spinal cord extracts in growth of neurites. Our results may be an important for the consideration of the hypothesis that ALS may be a disorder of motor neuron growth factors.     

Thyrotropin-releasing hormone (TRH): apparent receptor binding in rat spinal cord

Thyrotropin-releasing hormone (TRH) is unevenly distributed throughout the rat central nervous system including spinal cord, where exogenous TRH elicits profound pharmacological effects. [Pro-3H]TRH binds to 30,000 g pellet fraction from the spinal cord saturably, reversibly, and with high affinity (apparent Kd = 24-25 nM). This binding is displaced by TRH and related biologically active, but not inactive, peptides. TRH binding is evenly distributed throughout the rat spinal cord. Characteristics of this binding suggest an association with a physiologically relevant TRH receptor.     

Diffusible factor(s) from adult rat sciatic nerve increases cell number and neurite outgrowth of cultured embryonic ventral mesencephalic tyrosine hydroxylase-positive neurons

Dissociated embryonic rat ventral mesencephalon containing the developing A8-A10 dopamine (DA) neurons was cultured alone or in the presence of a 10 mm segment of adult rat sciatic nerve that had been explanted and maintained in separate culture for 72 hours prior to introduction to mesencephalic cultures. Nerve segments were contained in a co-culture basket, so that midbrain cells and nerve shared medium but were not in physical contact. The number and morphology of cultured DA neurons was assessed via immunocytochemistry for tyrosine hydroxylase (TH). Co-cultures of ventral midbrain tissue and nerve exhibited an increased number of TH-positive neurons, with larger neuronal perikarya, and increased length and complexity of neurites, than cultures of midbrain tissue alone. Increased number and growth of TH-positive neurons was obtained with as little as 2 days of exposure to nerve. This evidence suggests that a diffusible, soluble factor(s) from sciatic nerve can enhance the number and development of TH-positive neurons detected in cultures of embryonic ventral mesencephalon.     

Axon outgrowth from grafts of human embryonic spinal cord in the lesioned adult rat spinal cord.

Adult rats with acute partial lesions of their upper thoracic spinal cords were implanted bilaterally with cell suspensions of 6-7 week-old embryonic human spinal cord tissue one segment above or below the lesions. After 14-19 weeks, the animals were perfusion-fixed and the tissue analysed with a light microscope after immunocytochemical labelling with an antiserum recognizing human, but not rat, intermediary neurofilaments. Using this method, extensive efferent projections were demonstrated extending longitudinally from the grafts into the host spinal cord, both in the caudal and rostral directions. Within the white matter tracts, dense bundles of fibres extended for about 3-4 mm, and single fibres were identified up to 10 mm away from the implants. Axonal growth of this length within host white matter has not previously been observed from intraspinal grafts of rat CNS tissue.     

Trophic effect of angiotensin II, vasopressin and oxytocin on the ventral spinal cord of rat embryo

We studied trophic effects of angiotensin II, vasopressin and oxytocin on explanted ventral spinal cord cultures derived from 13 to 14-old day rat embryos. There was a significant neurite promoting effect in angiotensin II and vasopressin-treated cultures. Angiotensin II had the most potent effect at any concentrations. It became clear that minimum effective concentration was 10(-8)M in both angiotensin II and vasopressin. However, oxytocin had no neurotrophic effect at any concentrations. Our results demonstrated that angiotensin II and vasopressin have a neurotrophic effect on ventral spinal cord in cultures, and may contribute to therapeutic strategy of amyotrophic lateral sclerosis.     

Neurotrophic effect of angiotensin II, vasopressin and oxytocin on the ventral spinal cord of rat embryo

We studied trophic effects of angiotensin II, vasopressin and oxytocin on explanted ventral spinal cord cultures from 13-14-old day rat embryos. There was a significant neurite promoting effect in angiotensin II and vasopressin-treated cultures. Angiotensin II had the most potent effect at any concentrations. It became clear that minimum effective concentration was 10(-8) M in angiotensin II and vasopressin respectively. Effect of these two neuropeptides was concentration-dependent. However, oxytocin had no neurotrophic effect at any concentrations. Our results demonstrated that angiotensin II and vasopressin have a neurotrophic effect on ventral spinal cord in cultures, and may contribute to therapeutic strategy of amyotrophic lateral sclerosis.     

Receptors for thyrotropin-releasing hormone (TRH) in rabbit spinal cord

Specific binding of³H-labeled[3-Me-His²]TRH([³H]MeTRH) to membranes of rabbit spinal cord (thoracic) involved a homogeneous population of high-affinity sites (K_d = 2.7 ± 0.17 (n= 5)nM, B_max= 204 – 12(5)fmol/mg protein). TRH analogs competed for the binding in the following rank order of potency: MeTRH>TRH>TRH-Gly-NH₂ >Ser-His-Pro-NH₂ >Thr-His-Pro-NH₂ > pGlu-His-Pro-NH-C₂H₅>TRH-free acid. Competition experiments with rat amygdala, run in parallel with rabbit spinal cord, revealed a closely similar pattern of activity. These properties help identify binding sites in the rabbit spinal cord as physiological receptors for TRH. The binding sites resemble receptors previously demonstrated in pituitary and CNS tissues of other species.

The densities of [³H]MeTRH binding sites in different segments were generally similar, although density in the thoracic segment appeared to be somewhat higher. In all segments, binding seemed to be enriched in the dorsal gray matter. Dorsal roots and their associated ganglia, however, displayed little or no binding.     

Trophic effect of angiotensin II, vasopressin and other peptides on the cultured ventral spinal cord of rat embryo

We studied trophic effects of angiotensin II, vasopressin, cholecystokinin, and oxytocin on explanted ventral spinal cord cultures from 13- and 14-day-old rat embryos. There was a significant neurite promoting effect of the spinal cord cultures by using angiotensin II, vasopressin, and cholecystokinin. Cholecystokinin had the most potent effect at any concentrations. The minimum effective concentration was 10(-8) M in angiotensin II and vasopressin and 10(-12) M in cholecystokinin, respectively. The effect of angiotensin II and vasopressin was dependent on concentrations. However, the rate and grade of neurite appearance did not correlate with the concentrations of cholecystokinin. Oxytocin had no neurotrophic effect at any concentrations. Our results demonstrated that angiotensin II, vasopressin and cholecystokinin have neurotrophic effects on the ventral spinal cord in cultures, and may be candidates for therapeutic trials of amyotrophic lateral sclerosis.     

The development of a rat in vitro model of spinal cord injury demonstrating the additive effects of Rho and ROCK inhibitors on neurite outgrowth and myelination

It is currently thought that treatment for spinal cord injury (SCI) will involve a combined pharmacological and biological approach; however, testing their efficacy in animal models of SCI is time-consuming and requires large animal cohorts. For this reason we have modified our myelinating cultures as an in vitro model of SCI and studied its potential as a prescreen for combined therapeutics. This culture comprises dissociated rat embryonic spinal cord cells plated onto a monolayer of astrocytes, which form myelinated axons interspaced with nodes of Ranvier. After cutting the culture, an initial cell-free area appears persistently devoid of neurites, accompanied over time by many features of SCI, including demyelination and reduced neurite density adjacent to the lesion, and infiltration of microglia and reactive astrocytes into the lesioned area. We tested a range of concentrations of the Rho inhibitor C3 transferase (C3) and ROCK inhibitor Y27632 that have been shown to promote SCI repair in vivo. C3 promoted neurite extension into the lesion and enhanced neurite density in surrounding areas but failed to induce remyelination. In contrast, while Y27632 did not induce significant neurite outgrowth, myelination adjacent to the lesion was dramatically enhanced. The effects of the inhibitors were concentration-dependent. Combined treatment with C3 and Y27632 had additive affects with an enhancement of neurite outgrowth and increased myelination adjacent to the lesion, demonstrating neither conflicting nor synergistic effects when coadministered. Overall, these results demonstrate that this culture serves as a useful tool to study combined strategies that promote CNS repair.     

Effect of intrathecal proctolin administration on the behaviour evoked by the thyrotrophin-releasing hormone (TRH) analogue (RX 77368) and the indoleamine, TRH, substance P and calcitonin gene-related peptide levels and choline acetyltransferase activity in the rat spinal cord

The behavioral effects and the biochemical changes produced following either a single or repeated intrathecal injection respectively, of the insect peptide proctolin (Arg-Tyr-Leu-Pro-Thr-OH) have been compared with the effects of a stable analogue of thyrotrophin-releasing hormone (TRH) in rats. Intrathecal proctolin (1-100 micrograms) did not produce any marked behavioural effects on its own, while intrathecal TRH analogue (RX 77368, 0.5 microgram) administration produced wet-dog shakes and forepaw-licking behaviours. Proctolin (10 micrograms) significantly attenuated the wet-dog shake and forepaw-licking behaviours evoked by intrathecal RX 77368 administration when it was given 30 min before, but not when given in combination with RX 77368. Repeated intrathecal proctolin administration (10 micrograms twice daily for 5 days) significantly reduced the 5-hydroxytryptamine (5-HT), 5-hydroxyindoleacetic acid and TRH levels in the ventral, but not in the dorsal, horn of the spinal cord nor in the brainstem, and elevated hypothalamic TRH without affecting plasma free thyroxine levels when compared with values in saline-treated controls. Repeated proctolin injection did not alter substance P levels in any brain region examined, nor did it affect the choline acetyltransferase activity or the calcitonin gene-related peptide-like immunoreactive levels in the ventral horn of the spinal cord, both of which are principally located in motoneurones in this cord region.(ABSTRACT TRUNCATED AT 250 WORDS)