Activin receptor-like kinase 2 can mediate atrioventricular cushion transformation

The development of enteric and sympathetic neurons from neural crest precursor cells is regulated by signals produced by the embryonic environments to which the cells migrate. Bone morphogenetic proteins (BMPs) are present in the developing embryo and act to induce neuronal differentiation and noradrenergic properties of neural crest cells. We have investigated the role of BMP2 in regulating the appearance of distinct populations of autonomic neurons from postmigratory, HNK-1-positive neural crest precursor cells. BMP2 promotes neuronal differentiation of sympathetic and enteric precursor cells isolated from E14.5 rat. The effects of BMP2 change over time, resulting in a decrease in neuron number that can be attributed to apoptotic cell death. BMP2-dependent neuron death is rescued by gut-derived factors that provide trophic support to maturing neurons, indicating that BMP2 regulates the acquisition of trophic dependence of developing peripheral neurons. In addition to regulating neuron number, BMP2 promotes both panneuronal maturation and the acquisition of an enteric phenotype, as measured by lineage-specific changes in the expression of tyrosine hydroxylase and MASH-1. While BMP2 is sufficient to induce neuronal differentiation and panneuronal development, these results suggest that additional factors in the environment must collaborate with BMP2 to promote the final noradrenergic phenotype of sympathetic neurons.     

Huntingtin-interacting protein HIP14 is a palmitoyl transferase involved in palmitoylation and trafficking of multiple neuronal proteins

In neurons, posttranslational modification by palmitate regulates the trafficking and function of signaling molecules, neurotransmitter receptors, and associated synaptic scaffolding proteins. However, the enzymatic machinery involved in protein palmitoylation has remained elusive. Here, using biochemical assays, we show that huntingtin (htt) interacting protein, HIP14, is a neuronal palmitoyl transferase (PAT). HIP14 shows remarkable substrate specificity for neuronal proteins, including SNAP-25, PSD-95, GAD65, synaptotagmin I, and htt. Conversely, HIP14 is catalytically invariant toward paralemmin and synaptotagmin VII. Exogenous HIP14 enhances palmitoylation-dependent vesicular trafficking of several acylated proteins in both heterologous cells and neurons. Moreover, interference with endogenous expression of HIP14 reduces clustering of PSD-95 and GAD65 in neurons. These findings define HIP14 as a mammalian palmitoyl transferase involved in the palmitoylation and trafficking of multiple neuronal proteins.     

The development of the noradrenergic transmitter phenotype in postganglionic sympathetic neurons

Here we review recent data on molecular aspects of the differentiation of the noradrenergic neurotransmitter phenotype in postganglionic sympathetic neurons during avian and mammalian embryogenesis. By experimental manipulation of the chick embryo, it has been shown that neural tube and notochord are important for noradrenergic differentiation which occurs when migrating neural crest cells, the precursors of sympathetic ganglion cells, reach the dorsal aorta. Bone morphogenetic proteins expressed in the dorsal aorta before and during the time of noradrenergic differentiation are likely candidates for growth factors involved in induction of noradrenergic differentiation in vivo. To analyze noradrenergic differentiation, enzymes of the noradrenaline biosynthesis pathway and catecholamine stores have been used as differentiation markers. The molecules involved in neurotransmitter release which are as important for a functional noradrenergic neuron as those required for transmitter synthesis and storage are only recently being studied in this context. For a comprehensive view of the embryonic development of the noradrenergic neurotransmitter phenotype, it will be necessary to understand how the systems for synthesis, storage and release of noradrenaline are assembled during neuronal differentiation. Special issue dedicated to Dr. Hans Thoenen.     

Development of the cholinergic neurotransmitter phenotype in postganglionic sympathetic neurons

Sympathetic ganglia are composed of noradrenergic neurons and cholinergic neurons that differ in the expression of neurotransmitter-synthesizing enzymes, neurotransmitter transporters and neuropeptides. The analysis of the cholinergic differentiation during development revealed important principles involved in the generation of neuronal diversity, in particular the importance of signals from the innervated target. Some peripheral targets, such as the sweat glands in the mammalian footpads, are purely cholinergically innervated in the adult, whereas skeletal muscle arteries receive both noradrenergic and cholinergic innervation. For sympathetic neurons innervating sweat glands there is convincing evidence that these neurons are initially noradrenergic and that the interaction of innervating fibers and target tissue induces a shift in the neurotransmitter phenotype from noradrenergic to cholinergic. In addition to this target-dependent differentiation, an earlier expression of cholinergic characters was observed in sympathetic ganglia that occurs before target contact. These data raise the possibility that different subpopulations of cholinergic sympathetic neurons, innervating distinct peripheral targets, may develop along distinct schedules. In vitro studies suggest that growth factors of the family of neuropoietic cytokines are involved in the specification of the cholinergic sympathetic phenotype. Recent in vivo studies that interfered with cytokine receptor expression in developing avian sympathetic ganglia indicate that only the late, target-dependent differentiation depends on cytokine signaling. The signals involved in the early, target-independent expression of cholinergic properties remain to be determined, as well as the identity of the target-derived cytokine. Thus, cholinergic sympathetic differentiation seems to be more complex than expected, involving either both target-independent and target-dependent control or only target-induced differentiation, according to the specific neuronal subpopulation and target.     

Are mechanisms of exocytosis neurotransmitter specific?

In their review, Langley and Grant (1997) investigate the question whether mechanisms of exocytosis are neurotransmitter specific. There is now much evidence that the mechanisms governing the exocytosis of the two principal storage organelles—granules (large dense core vesicles) and electron-lucent vesicles—differ. But much less is known concerning potential differences in the release mechanisms of electron-lucent vesicles that store different types of fast neurotransmitters or of granules in different types of neurons. It is an open question whether there is a unifying control mechanism for the exocytosis of, for example, a peptide-containing granule of a glutamatergic neuron, a chromaffin granule, a noradrenergic granule or a granule from a neurosecretory neuron in the pituitary. The small electron-lucent synaptic vesicles of various kind apparently share common molecular components of regulated release. They carry the calcium sensor synaptotagmin, small GTP-binding proteins of the rab3 group or the v-SNARE synaptobrevin. Nevertheless, there may be differences in the regulatory mechanisms. This concerns the type of calcium channel involved or the absence of some of the presynaptic molecules such as rab3a, synapsin I or the t-SNAREs SNAP-25 or syntaxin from distinct types of neurons or sensory cells.     

The expression of dopamine beta-hydroxylase, tyrosine hydroxylase, and Phox2 transcription factors in sympathetic neurons: evidence for common regulation during noradrenergic induction and diverging regulation later in development

During differentiation of sympathetic neurons in chick embryos, tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH) mRNAs become detectable during the same developmental period and are both induced by BMP 4. Later during sympathetic ganglion development, DBH is detectable in TH-positive and -negative cells. Moreover, BMPs reduce DBH mRNA in cultures of sympathetic neurons while leaving TH unaffected. The data provide evidence for a common regulation of TH and DBH early during sympathetic neuron differentiation and indicate that BMPs promote their initial expression but not the maintenance during later development. The time course of Phox2a and 2b expression suggests an evolutionary conserved role in noradrenergic induction. In addition, Phox2a, Phox2b, and c-ret may be involved in the differentiation of cholinergic sympathetic neurons.     

Presynaptic trafficking of synaptotagmin I is regulated by protein palmitoylation

Protein palmitoylation plays a critical role in sorting and targeting of several proteins to pre- and postsynaptic sites. In this study, we have analyzed the role of palmitoylation in trafficking of synaptotagmin I and its modulation by synaptic activity. We found that palmitoylation of N-terminal cysteines contributed to sorting of synaptotagmin I to an intracellular vesicular compartment at the presynaptic terminal. Presynaptic targeting is a unique feature of N-terminal sequences of synaptotagmin I because the palmitoylated N terminus of synaptotagmin VII failed to localize to presynaptic sites. We also found that palmitate was stably associated with both synaptotagmin I and SNAP-25 and that rapid neuronal depolarization did not affect palmitate turnover on these proteins. However, long-term treatment with drugs that either block synaptic activity or disrupt SNARE complex assembly modulated palmitoylation and accumulation of synaptotagmin I at presynaptic sites. We conclude that palmitoylation is involved in trafficking of specific elements involved in transmitter release and that distinct mechanisms regulate addition and removal of palmitate on select neuronal proteins.     

BMP‐2 and cAMP elevation confer locus coeruleus neurons responsiveness to multiple neurotrophic factors

The locus coeruleus (LC) is a major target of several neurodegenerative disorders, including Parkinson's and Alzheimer's diseases. However, very little is known of the trophic requirements of LC neurons. In the present work, we have studied the biological activity of neurotrophic factors from different families in E15 primary cultures of LC neurons. In agreement with previous results, neurotrophin‐3 (NT‐3) and also glial cell line‐ derived neurotrophic factor (GDNF) increased the number of embryonic LC noradrenergic neurons in the presence of serum. In serum‐free conditions, none of the factors tested, including NT‐3, GDNF, neurturin, basic fibroblast growth factor (bFGF), or bone morphogenetic protein‐2 (BMP‐2), promoted the survival of tyrosine hydroxylase (TH)‐immunoreactive neurons at 6 days in vitro. However, when BMP‐2 was coadministered with any of these factors the number of LC TH‐positive neurons increased twofold. Similar results were obtained by cotreatment of LC neurons with forskolin and NT‐3, bFGF, or BMP‐2. The strongest effect (a fourfold increase in the number of TH‐positive cells) was induced by cotreatment with forskolin, BMP‐2, and GDNF. Thus, our results show that LC neurons require multiple factors for their survival and development, and suggest that activation of LC neurons by bone morphogenetic proteins and cAMP plays a decisive role in conferring noradrenergic neuron responsiveness to several trophic factors. © 2002 Wiley Periodicals, Inc. J Neurobiol 50: 291–304, 2002; DOI 10.1002/neu.10034     

Bone morphogenetic protein down-regulation of neuronal pituitary adenylate cyclase-activating polypeptide and reciprocal effects on vasoactive intestinal peptide expression

Among bone morphogenetic proteins (BMPs), the decapentaplegic (Dpp; BMP2, BMP4) and glass bottom boat (Gbb/60A; BMP5, BMP6, BMP7) subgroups have well-described functions guiding autonomic and sensory neuronal development, fiber formation and neurophenotypic identities. Evaluation of rat superior cervical ganglia (SCG) post-ganglionic sympathetic neuron developmental regulators identified that selected BMPs of the transforming growth factor beta superfamily have reciprocal effects on neuronal pituitary adenylate cyclase-activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP) expression. Dpp and Gbb/60A BMPs rapidly down-regulated PACAP expression, while up-regulating other sympathetic neuropeptides, including PACAP-related VIP. The suppressive effects of BMP on PACAP mRNA and peptide expression were potent, efficacious and phosphorylated mothers against decapentaplegic homolog (Smad) signaling-dependent. Axotomy of SCG dramatically increases PACAP expression, and the possibility that abrogation of inhibitory retrograde target tissue BMP signaling may contribute to this up-regulation of sympathetic neuron PACAP was investigated. Replacement of BMP6 to SCG explant preparations significantly blunted the injury-induced elevated PACAP expression, with a concomitant decrease in sympathetic PACAP-immunoreactive neuron numbers. These studies suggested that BMPs modulate neuropeptide identity and diversity by stimulating or restricting the expression of specific peptidergic systems. Furthermore, the liberation of SCG neurons from target-derived BMP inhibition following axotomy may be one participating mechanism associated with injury-induced neuropeptidergic plasticity.