Synaptic development in the crayfish opener muscle

Nerve terminal regions in walking leg opener muscles of several crayfish of different ages (0 to 245 days after hatching) were examined by means of electron microscopy. This muscle is innervated by two axons (excitatory and inhibitory) and at maturity contains three classes of synapse: excitatory and inhibitory neuromuscular synapses, and inhibitory axo-axonal synapses. The muscle itself is initially a syncytium, which gradually becomes subdivided into distinct "muscle fibers" as the animal matures. Innervation was not found in the opener muscle just before or just after hatching, but was present in restricted locations on the inner side of the muscle within a few days of hatching. As the muscle enlarged and became subdivided, innervation appeared in various other locations. Synaptic contacts were located in young stages soon after hatching, and in later stages. Morphological differences characteristic of excitatory nerve terminals could be found even at the earliest stages of innervation. Both excitatory and inhibitory synapses, but particularly the former, showed evidence of progressive enlargement to a final size within the first two months, and no evidence for further enlargement of existing synapses thereafter. Synaptic maturation also involved the appearance of presynaptic "dense bodies" though to be regions at which transmitter substance is preferentially released. Nerve terminals at different levels of maturation were observed in opener muscles of young crayfish. Clear evidence for differential maturation of the three types of synapse present in this muscle was obtained. The inhibitory neuromuscular synapses attained their final average size and developed their dense bodies sooner than the excitatory neuromuscular synapses. The inhibitory axo-axonal synapses were the last to appear and to mature.     

BEN/SC1/DM-GRASP expression during neuromuscular development: a cell adhesion molecule regulated by innervation

BEN/SC1/DM-GRASP is a cell adhesion molecule belonging to the Ig superfamily that is transiently expressed during avian embryogenesis in a variety of cell types, including the motoneurons of the spinal cord. We have investigated the pattern of BEN expression during neuromuscular development of the chick. We show that both motoneurons and their target myoblasts express BEN during early embryonic development and that the protein becomes restricted at neuromuscular contacts as soon as postsynaptic acetylcholine receptor clusters are observed in muscle fibers. Muscle cells grown in vitro express and maintain BEN expression even when they fuse and give rise to mature myotubes. When embryos are deprived of innervation by neural tube ablation, BEN expression is observed in muscle fibers, whereas, in control, the protein is already restricted at neuromuscular synaptic sites. These results demonstrate that all myogenic cells intrinsically express BEN and maintain the protein in the absence of innervation. Conversely, when neurons are added to myogenic cultures, BEN is rapidly downregulated in muscle cells, demonstrating that innervation controls the restricted pattern of BEN expression seen in innervated muscles. After nerve section in postnatal muscles, BEN protein becomes again widely spread over muscle fibers. When denervated muscles are allowed to be reinnervated, the protein is reexpressed in regenerating motor axons, and reinnervation of synaptic sites leads to the concentration of BEN at neuromuscular junctions. Our results suggest that BEN cell adhesion molecule acts both in the formation of neuromuscular contacts during development and in the events leading to muscle reinnervation.     

Fusimotor-free spindles in reinnervated muscles of neonatal rats treated with nerve growth factor

Crushing the nerve to the medial gastrocnemius muscle in newborn rats and administering nerve growth factor afterwards results in a reinnervated muscle containing supernumerary muscle spindles. The structure and innervation of 88 spindles in the reinnervated muscles were reconstructed from serial thick and thin transverse sections at 30-35 days after the nerve crush, and compared to those of five control spindles. The spindles consisted of one to four small-diameter encapsulated fibers with features of nuclear chain intrafusal fibers, or infrequently a nuclear bag intrafusal fiber. Some of the spindles were located within a capsule that also contained an extrafusal fiber. Each spindle was innervated by an afferent with features of the primary afferent. The density of secondary afferents was lower in reinnervated muscles than in controls. Endplates were observed on extrafusal fibers in the experimental muscles, attesting to restoration of skeletomotor (alpha) innervation after the nerve crush. However, 78% of the experimental spindles were entirely devoid of efferent innervation. The remainder received either one or two fusimotor (gamma) axons or a skeletofusimotor (beta) axon, compared to the six to eight motor axons that innervated control spindles. The presence of supernumerary spindles composed of fibers that resemble normal intrafusal fibers in the absence of motor innervation suggests that afferents alone can induce the formation and subsequent differentiation of intrafusal fibers in nerve-crushed muscles of neonatal rats. In addition, the paucity of gamma innervation in nerve-crushed muscles suggests that immature gamma neurons are more susceptible than spindle afferents or alpha efferents to cell death after axotomy at birth.     

Specificity and development of innervation pattern of Xenopus pectoralis muscle

Multielectrode recordings were used to identify and measure the axonal inputs to each end plate on contiguous surface fibers covering about 25% of the Xenopus pectoralis muscle in mature and developing animals. The mature innervation pattern was remarkably precise. Individual axons tended to innervate fibers of similar input resistance (R(in)) in compact motor units restricted to only a portion of the region studied. Motor units comprising fibers of similar R(in) overlapped mainly near their borders. Most fibers had two end plates. In more than 80% of these fibers, both end plates received input from the same axon. In 57%, this was the only input to both end plates. This implies a powerful mechanism for excluding or eliminating inputs from other axons. About 16% of the mature junctions showed focal polyneuronal innervation, with the weaker end plate potential component often less than 1 mV in noncurarized preparation. However, we have no evidence that the weaker inputs were being eliminated. During development, motor units became more compact, which was associated with synapse elimination; but from the earliest times studied, soon after metamorphosis when many fibers were adding second end plates, a majority of those that had two end plates were innervated at both sites by the same axon.     

Cell death of spinal motoneurons in the chick embryo following deafferentation: rescue effects of tissue extracts, soluble proteins, and neurotrophic agents

In the absence of descending spinal and supraspinal afferent inputs, neurons in the developing lumbar spinal cord of the chick embryo undergo regressive changes including cellular atrophy and degeneration between embryonic days 10 and 16. There are significant decreases in the number of motoneurons, interneurons, and sensory (dorsal root ganglion) neurons. Although there are several possible explanations for how afferents might regulate the maintenance of neuronal viability, we have focused attention on the putative role of neurotrophic agents in these events. Previous studies have shown that specific tissue extracts (e.g., muscle, brain), soluble proteins, growth factors, and trophic agents can promote the in vitro and in vivo survival of avian motoneurons during the period of natural cell death (embryonic days 6-10). Several of these agents were also effective following deafferentation. These included brain extract (BEX), muscle extract (MEX), conditioned medium from astrocyte cultures (ACM), as well as the following neurotrophic agents: nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3), S-100, insulin-like growth factor-I (IGF-I), ciliary neurotrophic factor (CNTF), platelet-derived growth factor (PDGF), basic fibroblast growth factor (bFGF), and leukemia inhibitory factor (CDF/LIF). Both transforming growth factor-beta (TGF-beta) and acidic fibroblast growth factor (aFGF) were ineffective. Although considerable more work is needed to determine which (and how) specific CNS-derived trophic agents regulate motoneuron survival, the present results are consistent with the notion that neurotrophic agents released from or modulated by synaptic inputs to target neurons promote neuronal differentiation and survival in the CNS.     

Skeletal muscle extract and nerve growth factor have developmentally regulated survival promoting effects on distinct populations of mammalian sensory neurons

Neurotrophic factors appear to be relevant to the therapy of degenerative diseases as well as neural regeneration. In this respect, we have investigated the neurotrophic effects of skeletal muscle extract on DRG neuron survival by examining the survival and neurite outgrowth promoting activity of factor(s) present in skeletal muscle extracts (SME) on dissociated cultures of embryonic or early postnatal mouse dorsal root ganglion (DRG) sensory neurons. The numbers of surviving neurons resulting from SME addition increased continuously from embryonic day 13 (15%) to birth (55%), then decreased up to 7 days after hatching (0%). Preliminary characterization of the factor(s) present in SME suggests that the active molecule is a protein different from the known neurotrophic factors NGF, BDNF, NT3, CNTF, and bFGF, and that its neurotrophic effect is not mediated by direct interaction with the substratum.     

Differential effects of a six-day immobilization on newborn rat soleus muscles at two developmental stages

Our objective was to determine the effects of a six-day immobilization on the musculoskeletal system of the rat during postnatal development at two key periods when the states of innervation are known to be different. This work was undertaken on the soleus muscle since it is well known that postural slow muscles show marked changes after a period of disuse. Thus, the soleus muscle was immobilized in a shortened position either when the innervation was polyneuronal or monosynaptic, respectively from 6 to 12 and from 17 to 23 days. The muscle modifications were followed by ATPase staining and myosin heavy chain (MyHC) isoform identification using monoclonal antibodies and SDS-PAGE. The functional properties of skinned fibre bundles were established by calcium/strontium (Ca/Sr) activation characteristics. In control muscles the maturation was characterized by a progressive increase of adult MyHCs (I and IIA) concomitant with a decrease in both the MyHC neo and the Ca affinity. Between 6 to 12 days, immobilization of the limb induced an increase in histochemical type IIC fibres. Using antibodies we identified new fibre types, classified as a function of their MyHC isoform co-expression. We observed an increase in expression of both MyHC neo and Ca affinity. From 17 to 23 days, the immobilization induced an increase in Ca affinity and marked changes in the MyHC isoform composition: disappearance of MyHC neo and expression of the fast MyHC IIB isoform, which in normal conditions is never expressed in the soleus muscle. We conclude that an immobilization imposed during polyneuronal innervation delays the postnatal maturation of the soleus muscle, whereas when the immobilization is performed under monosynaptic innervation the muscle evolves towards a fast phenotype using a default pathway for MyHC expression.     

Morphological aspects of the elimination of polyneuronal innervation of skeletal muscle fibres in newborn rats

The ultrastructure of neuromuscular junctions of rat soleus muscles 1-40 days postnatally was examined for possible morphological correlates of the transient polyneuronal innervation which is present in newborn rats. Several vesicle-laden profiles of terminal axons are seen to contact each muscle fiber up to 8 days postnatally. Axon terminals often lie close together, without Schwann cell intervention. Between days 8 and 16 the number of profiles of terminals on each muscle fibre is reduced, and both Schwann cells and ridge-like extensions of muscle fibre cytoplasm intervene between and separate axon terminals. No signs of degenerating intramuscular axons or axon terminals could be found. It is suggested that the redundant terminals are eliminated by retraction into the parent axons. This process is apparently accomplished without any morphological signs of degeneration.     

Developmental regulation of the serpin, protease nexin I, localization during activity-dependent polyneuronal synapse elimination in mouse skeletal muscle

During vertebrate neuromuscular development, all muscle fibers are transiently innervated by more than one neuron. Among the numerous factors shown to potentially influence the passage from poly- to mononeuronal innervation, serine proteases and their inhibitors appear to play important roles. In this regard, protease nexin I (PNI), a potent inhibitor of the serine protease, thrombin, is highly localized to the neuromuscular junction (NMJ). In turn, thrombin is responsible for activity-dependent synapse elimination both in an in vitro model, and in vivo. In the present study, we used a monospecific anti-PNI polyclonal antibody to study the developmental kinetics of PNI expression in mouse leg skeletal muscle. By using immunoblotting, we detected PNI at embryonic day 16 (E16), as a 48-kDa band. This 48-kDa PNI band became prominent in leg muscle extracts at postnatal day 5 (P5) and remained so in extracts from adult muscle. In contrast, a higher molecular weight immunoreactive PNI band, which was sodium dodecyl sulfate- and beta-mercaptoethanol-resistant, was first detected at E16, increased at birth (P0), and then decreased at P15, i.e., after the wave of polyneuronal synapse elimination had occurred in these muscles. The results of an enzyme-linked immunosorbent assay, measuring active, complexed, and truncated PNI, correlated with Western blot data. We used immunocytochemistry to probe the localization of PNI at the NMJ and found that PNI was present in the cytoplasm of myotubes at E16, but neither then nor at birth did it colocalize with acetylcholine receptors. PNI became localized at NMJs by P5 and increased by P15, after which it remained stably concentrated there in the adult. Finally, we studied the gene expression of PNI mRNA, by using Northern blotting, and showed that PNI mRNA was present in skeletal muscle and remained stable throughout the time-course studies, suggesting that developmental regulation of muscle PNI occurs principally at the translational and/or post-translational levels. These results suggest that the localization of PNI, through a binding site or "receptor" may play an important role in differentiation and maintenance of synapse.     

Basic fibroblast growth factor enhances the growth of postnatal neostriatal GABAergic neurons in vitro

Basic fibroblast growth factor (bFGF) significantly enhances the short-term survival of embryonic striatal neurons in vitro but has little effect on the outgrowth of striatal cells compared to neurons from other brain regions. Studies in our laboratory have shown that bFGF protects postnatal striatal cells in vitro from NMDA receptor-induced neurotoxicity. We therefore examined the effects of bFGF on the outgrowth of GABA-containing cells taken from the postnatal (Day 1) caudate-putamen and cultured for up to 3 weeks. In control cultures GABAergic neurons formed three populations based on somatic size and developed the cytoarchitectural features characteristic of dendrites, spines, and axons. In the presence of bFGF (6 pM continuously from the day of plating), small- and medium-sized GABAergic neurons showed significant increases compared to untreated controls in axon-like growth (axon length) at 6 days in culture and in both axon- and dendrite-like neurite growth (axon length and branch order, number of primary dendrites, dendrite length, and dendritic branch order) at 13 and 17 days in culture. Large GABAergic neurons were unaffected by treatment with bFGF. Striatal GABAergic neurons exposed to nerve growth factor (10 ng/ml) were not different from untreated controls. Neuron survival was also unaffected by bFGF treatment at all days in culture examined. Other observations suggested that the neurotrophic effects of bFGF were mediated by a direct action of the growth factor on striatal neurons and not glial cells. First, glial cells (identified by the immunohistochemical localization of glial fibrillary acidic protein) were unaffected by bFGF treatment at the low concentration (6 pM) used to enhance neurite growth, but did significantly proliferate at higher concentrations of bFGF (6 nM). Second, immunoreactive bFGF receptor protein was localized predominantly to the somata and processes of striatal neurons and not to glial cells in the cultures. Finally, when neurons from control cultures were briefly exposed (1 to 4 h) to bFGF at concentrations which were neurotrophic, a marked elevation in the immediate early gene protein c-fos was observed by immunohistochemistry in the nuclei of neurons, including GABAergic cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

Systemic injections of ciliary neurotrophic factor induce sprouting by adult motor neurons

The ability of ciliary neurotrophic factor (CNTF) to induce sprouting by undamaged adult motor neurons was studied in gluteal muscles of adult ICR mice. Low doses of CNTF (0.02 mg kg-1 day-1) only induced sprouting in gluteus muscles that were beneath the site of injection, whereas high doses of CNTF (0.4-1.2 mg kg-1 day-1) acted systemically to induce motor neuron sprouting. We found little difference between the type or quality of sprouting induced by CNTF compared with muscle paralysis. Both stimuli induced sprouts of the same length, although muscle paralysis tended to induce more sprouts per end-plate. Paralysis also induced more nodal sprouting than did CNTF, but both were weaker stimuli for nodal sprouting than was partial denervation. In addition to its effects on motor neuron sprouting, high doses of CNTF induced loss of up to 36% of the body weight of treated mice. The substantial wasting caused by CNTF indicates that the factor has potent cachectic activity.     

Induction of basic fibroblast growth factor mRNA by basic fibroblast growth factor in Müller cells

PURPOSE: To investigate the induction of basic fibroblast growth factor (bFGF) gene expression in cultured rat Müller cells by bFGF and to study the mechanism of induction. METHODS: Müller cells from 1- to 3-day-old Sprague-Dawley rats were isolated and cultured with Dulbecco's modified Eagle's medium with 10% fetal calf serum. Cultured cells were identified by immunocytochemistry using antibodies against vimentin, carbonic anhydrase II, and glutamine synthetase. Cells of passages 1 through 4 were treated with bFGF, the protein kinase C (PKC) inhibitor, H-7; calphostin C, or the PKC activator, PMA; and protein kinase A (PKA) inhibitor, H-89; as well as the adenylate cylase activator, forskolin; or the adenylate cyclase inhibitor, SQ22536. Northern blot analysis was performed to determine the mRNA expression of bFGF, ciliary neurotrophic factor (CNTF) and brain-derived neurotrophic factor (BDNF). RESULTS: Addition of bFGF to culture medium induced bFGF gene expression in a dose- and time-dependent manner. Induction of bFCF mRNA started at a bFGF concentration of 0.1 ng/ml. The bFGF mRNA level was elevated by 2-fold at 1 ng/ml of bFGF, 2.8-fold at 5 ng/ml, and reached a peak of 4-fold at 10 ng/ml and 3.7-fold at 50 ng/ml. At 10 ng/ml of bFGF, induction of bFGF mRNA was observed as early as 2 hours (2-fold) after treatment. The bFGF mRNA level continued to increase to 3.7-fold by 4 hours, and reached a maximum of 4.4-fold by 8 hours. A slow decline of the bFGF mRNA level was observed after 8 hours of bFGF treatment (3.5-fold by 12 hours, and 3-fold by 24 hours). This induction of bFGF gene expression was blocked by PKC inhibitors H-7 (30 microM). The PKC activator PMA (0.1 microM) also upregulated bFGF gene expression, but the effects of bFGF and PMA were not additive. An adenylate cyclase inhibitor, SQ22536 (100 microM), did not inhibit bFGF-induced bFGF gene expression. Although forskolin (5 microM), an adenylate cyclase activator, also upregulated the level of bFGF mRNA, the effects of forskolin and bFGF were additive. In addition, no inhibitory effect on bFGF-induced expression of bFGF mRNA was found using H-89 (1 microM). Exogenous bFGF did not alter the mRNA levels of CNTF and BDNF. CONCLUSIONS: These results indicate that bFGF induces bFGF gene expression in cultured rat Müller cells through PKC activation. The authors' findings raise the possibility that Müller cells in vivo also respond to available bFGF (for example, that released from the endogenous reservoirs in the case of injury) or to exogenous bFGF by producing more bFGF, which could in turn promote photoreceptor survival.     

Laser ablation of Drosophila embryonic motoneurons causes ectopic innervation of target muscle fibers

We have tested the effects of neuromuscular denervation in Drosophila by laser-ablating the RP motoneurons in intact embryos before synaptogenesis. We examined the consequences of this ablation on local synaptic connectivity in both 1st and 3rd instar larvae. We find that the partial or complete loss of native innervation correlates with the appearance of alternate inputs from neighboring motor endings and axons. These collateral inputs are found at ectopic sites on the denervated target muscle fibers. The foreign motor endings are electrophysiologically functional and are observed on the denervated muscle fibers by the 1st instar larval stage. Our data are consistent with the existence of a local signal from the target environment, which is regulated by innervation and influences synaptic connectivity. Our results show that, despite the stereotypy of Drosophila neuromuscular connections, denervation can induce local changes in connectivity in wild-type Drosophila, suggesting that mechanisms of synaptic plasticity may also be involved in normal Drosophila neuromuscular development.     

Schwann cells proliferate at rat neuromuscular junctions during development and regeneration

Terminal Schwann cells (TSCs) cover neuromuscular junctions and are important in the repair and maintenance of these synapses. We have examined how these cells are generated at developing junctions and how their number is regulated during repair of nerve injury. At birth, approximately half of the junctions in rat soleus and extensor digitorum longus muscles have one TSC soma. Somata are absent from the remainder, although Schwann cell (SC) processes arising from somata along the preterminal axon cover almost all of these synapses. By 2 months of age, junctions have gained an additional two to three TSCs. Most of this gain occurs during the first 2 postnatal weeks and largely precedes the expansion of endplate size. Although the initial addition is caused by cell migration, mitotic labeling shows extensive division of TSCs at junctions. A slower addition of TSCs occurs in adult muscles, and TSC number in the adult is correlated with endplate size. During repair of nerve injury, TSC number is regulated by a combination of signals from motor neurons and denervated tissue. As shown previously (Connor et al., 1987), denervation of adult muscles did not, in itself, cause TSC mitosis. However, TSCs became mitotic during reinnervation. Partial denervation induced division of TSCs at innervated but not denervated endplates. A disproportionate number of these mitotic cells were found at endplates contacted by TSC processes extended from nearby denervated endplates, contacts known to promote nerve sprouting. These results show an association between TSC mitotic activity and alterations in synaptic structure during development, sprouting, and reinnervation.     

Glucocorticoid facilitation of cholinergic development in the rat hippocampus

The role of endogenous glucocorticoids in facilitating the postnatal innervation of septohippocampal cholinergic projections was examined. Septohippocampal cholinergic innervation was determined using two methods. One method involved measuring the optical density of acetylcholinesterase, a marker of cholinergic fibres in the hippocampus. In the other method, acetylcholinesterase-positive fibre counts were made in the hippocampus. Both methods revealed that 14-day-old rats adrenalectomized at 10 days of age have significantly lower densities of acetylcholinesterase in the hippocampal dentate gyrus molecular layer and in the regio inferior when compared to sham-operated control rats. This reduction in hippocampal acetylcholinesterase did not occur when 10-day-old adrenalectomized rats were either injected daily with exogenous corticosterone (0.3 mg/100 g body weight) or when adrenalectomy was conducted at later postnatal ages. In addition, unlike the developing hippocampus, the basolateral nucleus of the amygdala, which is also highly innervated by cholinergic fibres, showed no significant changes in acetylcholinesterase density after adrenalectomy. These observations suggest that glucocorticoids play an important role in supporting the development of cholinergic projections to the hippocampus. Cholinergic innervation of the hippocampus appears especially sensitive to the action of glucocorticoids occurring before the conclusion of the second postnatal week. Furthermore, this glucocorticoid influence is directed rather specifically to the hippocampus in comparison to the basolateral amygdala.     

In vitro survival and differentiation of neurons derived from epidermal growth factor-responsive postnatal hippocampal stem cells: inducing effects of brain-derived neurotrophic factor

Neural stem cells proliferate in vitro and form neurospheres in the presence of epidermal growth factor (EGF), and are capable of differentiating into both neurons and glia when exposed to a substrate. We hypothesize that specific neurotrophic factors induce differentiation of stem cells from different central nervous system (CNS) regions into particular fates. We investigated differentiation of stem cells from the postnatal mouse hippocampus in culture using the following trophic factors (20 ng/mL): brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and glial-derived neurotrophic factor (GDNF). Without trophic factors, 32% of stem cells differentiated into neurons by 4 days in vitro (DIV), decreasing to 10% by 14 DIV. Addition of BDNF (starting at either day 0 or day 3) significantly increased neuron survival (31-43% by 14 DIV) and differentiation. Morphologically, many well-differentiated neurons resembled hippocampal pyramidal neurons. 5'-Bromodeoxyuridine labeling demonstrated that the pyramidal-like neurons originated from stem cells which had proliferated in EGF-containing cultures. However, similar application of NT-3 and GDNF did not exert such a differentiating effect. Addition of BDNF to stem cells from the postnatal cerebellum, midbrain, and striatum did not induce these neuronal phenotypes, though similar application to cortical stem cells yielded pyramidal-like neurons. Thus, BDNF supports survival of hippocampal stem cell-derived neurons and also can induce differentiation of these cells into pyramidal-like neurons. The presence of pyramidal neurons in BDNF-treated hippocampal and cortical stem cell cultures, but not in striatal, cerebellar, and midbrain stem cell cultures, suggests that stem cells from different CNS regions differentiate into region-specific phenotypic neurons when stimulated with an appropriate neurotrophic factor.     

Keratinocyte gene transfer and gene therapy

Ciliary neurotrophic factor (CNTF) is a multifunctional cytokine that mediates survival and differentiation of neurons as well as many other cell types. In this study, CNTF and leukemia inhibitory factor (LIF) reduced the apparent number of primary serotonergic neurons in E14 raphe culture by 90% as determined by immunocytochemistry for serotonin (5HT). The reduction in 5HT cell number was not due to neuronal loss as removal of CNTF after 4 days in culture resulted in a partial restitution of the serotonergic phenotype. In the RN46A serotonergic cell line which is induced to become serotonergic by brain-derived neurotrophic factor (BDNF), the addition of CNTF suppressed tryptophan hydroxylase and 5HT synthesis and increased choline acetyl transferase (ChAT) expression by 6-fold and ChAT activity by 20- to 30-fold over 12 days. As with the primary neurons, removal and replacement of CNTF with BDNF after 4 days resulted in a partial restitution of 5HT expression. Moreover, other members of the CNTF-cytokine family that use gp130 and/or LIF receptor beta as their signal transducing receptors-LIF, oncostatin M, interleukin 6, and interleukin 11-had similar effects on increasing ChAT activity and reducing 5HT expression in RN46A cells. Analysis of 5HT levels showed no significant difference in the amount of serotonin between wild-type and CNTFR alpha knockout mice at birth, suggesting that the potential to switch phenotype mediated through CNTFR alpha is a latent property of neuroepithelial precursors in the raphe nucleus.     

Fc gamma RIIIB gene duplication: evidence for presence and expression of three distinct Fc gamma RIIIB genes in NA(1+,2+)SH(+) individuals

To investigate when the neurotrophic cytokines ciliary neurotrophic factor (CNTF), leukaemia inhibitory factor (LIF), oncostatin-M (OSM), interleukin-6 (IL-6) and cardiotrophin-1 (CT-1) act on developing sensory neurones and whether they co-operate with neurotrophins in regulating neuronal survival, we studied the in vitro trophic effects of these factors on two well-characterized populations of cranial sensory neurones at closely staged intervals throughout embryonic development. The cutaneous sensory neurones of the trigeminal ganglion, which show an early, transient survival response to BDNF and NT3 before becoming NGF-dependent, were supported by CNTF, LIF, OSM and CT-1 during the late fetal period, several days after the neurones become NGF-dependent. At this stage of development, these cytokines promoted the survival of a subset of NGF-responsive neurones. The enteroceptive neurones of the nodose ganglion, which retain dependence on BDNF throughout fetal development, were supported throughout their development by CNTF, LIF, OSM and CT-1, and displayed an additional survival response to IL-6 in the late fetal period. These findings indicate that populations of sensory neurones display different developmental patterns of cytokine responsiveness and show that embryonic trigeminal neurones pass through several phases of differing neurotrophic factor survival requirements.