FGF8 induces formation of an ectopic isthmic organizer and isthmocerebellar development via a repressive effect on Otx2 expression. (1/1984)

Beads containing recombinant FGF8 (FGF8-beads) were implanted in the prospective caudal diencephalon or midbrain of chick embryos at stages 9-12. This induced the neuroepithelium rostral and caudal to the FGF8-bead to form two ectopic, mirror-image midbrains. Furthermore, cells in direct contact with the bead formed an outgrowth that protruded laterally from the neural tube. Tissue within such lateral outgrowths developed proximally into isthmic nuclei and distally into a cerebellum-like structure. These morphogenetic effects were apparently due to FGF8-mediated changes in gene expression in the vicinity of the bead, including a repressive effect on Otx2 and an inductive effect on En1, Fgf8 and Wnt1 expression. The ectopic Fgf8 and Wnt1 expression domains formed nearly complete concentric rings around the FGF8-bead, with the Wnt1 ring outermost. These observations suggest that FGF8 induces the formation of a ring-like ectopic signaling center (organizer) in the lateral wall of the brain, similar to the one that normally encircles the neural tube at the isthmic constriction, which is located at the boundary between the prospective midbrain and hindbrain. This ectopic isthmic organizer apparently sends long-range patterning signals both rostrally and caudally, resulting in the development of the two ectopic midbrains. Interestingly, our data suggest that these inductive signals spread readily in a caudal direction, but are inhibited from spreading rostrally across diencephalic neuromere boundaries. These results provide insights into the mechanism by which FGF8 induces an ectopic organizer and suggest that a negative feedback loop between Fgf8 and Otx2 plays a key role in patterning the midbrain and anterior hindbrain.  (+info)

N-type voltage-dependent calcium channels mediate the nicotinic enhancement of GABA release in chick brain. (2/1984)

The role of voltage-dependent calcium channels (VDCCs) in the nicotinic acetylcholine receptor (nAChR)-mediated enhancement of spontaneous GABAergic inhibitory postsynaptic currents (IPSCs) was investigated in chick brain slices. Whole cell recordings of neurons in the lateral spiriform (SpL) and ventral lateral geniculate (LGNv) nuclei showed that cadmium chloride (CdCl2) blocked the carbachol-induced increase of spontaneous GABAergic IPSCs, indicating that VDCCs might be involved. To conclusively show a role for VDCCs, the presynaptic effect of carbachol on SpL and LGNv neurons was examined in the presence of selective blockers of VDCC subtypes. omega-Conotoxin GVIA, a selective antagonist of N-type channels, significantly reduced the nAChR-mediated enhancement of gamma-aminobutyric acid (GABA) release in the SpL by 78% compared with control responses. Nifedipine, an L-type channel blocker, and omega-Agatoxin-TK, a P/Q-type channel blocker, did not inhibit the enhancement of GABAergic IPSCs. In the LGNv, omega-Conotoxin GVIA also significantly reduced the nAChR-mediated enhancement of GABA release by 71% from control values. Although omega-Agatoxin-TK did not block the nicotinic enhancement, L-type channel blockers showed complex effects on the nAChR-mediated enhancement. These results indicate that the nAChR-mediated enhancement of spontaneous GABAergic IPSCs requires activation of N-type channels in both the SpL and LGNv.  (+info)

Midbrain combinatorial code for temporal and spectral information in concurrent acoustic signals. (3/1984)

All vocal species, including humans, often encounter simultaneous (concurrent) vocal signals from conspecifics. To segregate concurrent signals, the auditory system must extract information regarding the individual signals from their summed waveforms. During the breeding season, nesting male midshipman fish (Porichthys notatus) congregate in localized regions of the intertidal zone and produce long-duration (>1 min), multi-harmonic signals ("hums") during courtship of females. The hums of neighboring males often overlap, resulting in acoustic beats with amplitude and phase modulations at the difference frequencies (dFs) between their fundamental frequencies (F0s) and harmonic components. Behavioral studies also show that midshipman can localize a single hum-like tone when presented with a choice between two concurrent tones that originate from separate speakers. A previous study of the neural mechanisms underlying the segregation of concurrent signals demonstrated that midbrain neurons temporally encode a beat's dF through spike synchronization; however, spectral information about at least one of the beat's components is also required for signal segregation. Here we examine the encoding of spectral differences in beat signals by midbrain neurons. The results show that, although the spike rate responses of many neurons are sensitive to the spectral composition of a beat, virtually all midbrain units can encode information about differences in the spectral composition of beat stimuli via their interspike intervals (ISIs) with an equal distribution of ISI spectral sensitivity across the behaviorally relevant dFs. Together, temporal encoding in the midbrain of dF information through spike synchronization and of spectral information through ISI could permit the segregation of concurrent vocal signals.  (+info)

NMDA receptor characterization and subunit expression in rat cultured mesencephalic neurones. (4/1984)

1. NMDA-induced changes in free intracellular Ca2+ concentration ([Ca2+]i) were determined in individual cultured rat mesencephalic neurones by the fura-2 method. mRNA expression encoding NMDA receptor subunits (NR1, NR2A-D) was examined by RT-PCR. 2. NMDA (1-100 microM, plus 10 microM glycine) induced a concentration-dependent increase in [Ca2+]i (EC50 = 5.7 microM). The effect of NMDA was virtually insensitive to tetrodotoxin (0.3 microM) and nitrendipine (1 microM), but dependent on extracellular Ca2+. 5,7-Dichlorokynurenic acid (10 microM), a specific antagonist at the glycine binding site on the NMDA receptor, abolished the NMDA response. 3. Memantine, an open-channel blocker, and ifenprodil, a preferential non-competitive NR1/NR2B receptor antagonist diminished the NMDA effect with an IC50 value of 0.17 and 1 microM, respectively. Ethanol at 50 and 100 mM caused about 25 and 45%-inhibition, respectively. 4. Agarose gel analysis of the PCR products followed by ethidium bromide fluorescence or CSPD chemiluminescence detection revealed an almost exclusive expression of the NR1 splice variants lacking exon (E) 5 and E22. The 3' splice form without both E21 and E22 exceeded that containing E21 by approximately 4 fold. The relative amounts of NR2A, NR2B, NR2C corresponded to approximately 1:2:1. NR2D mRNA was also detectable. 5. In conclusion, mesencephalic neurones bear ethanol-sensitive NMDA receptors which might be involved in the development of ethanol dependence and withdrawal. The high affinity of NMDA to this receptor, its sensitivity to ifenprodil and memantine may suggest that the mesencephalic NMDA receptor comprises the NR1 splice variant lacking E5, NR2B, and NR2C, respectively.  (+info)

A binding site for homeodomain and Pax proteins is necessary for L1 cell adhesion molecule gene expression by Pax-6 and bone morphogenetic proteins. (5/1984)

The cell adhesion molecule L1 regulates axonal guidance and fasciculation during development. We previously identified the regulatory region of the L1 gene and showed that it was sufficient for establishing the neural pattern of L1 expression in transgenic mice. In the present study, we characterize a DNA element within this region called the HPD that contains binding motifs for both homeodomain and Pax proteins and responds to signals from bone morphogenetic proteins (BMPs). An ATTA sequence within the core of the HPD was required for binding to the homeodomain protein Barx2 while a separate paired domain recognition motif was necessary for binding to Pax-6. In cellular transfection experiments, L1-luciferase reporter constructs containing the HPD were activated an average of 4-fold by Pax-6 in N2A cells and 5-fold by BMP-2 and BMP-4 in Ng108 cells. Both of these responses were eliminated on deletion of the HPD from L1 constructs. In transgenic mice, deletion of the HPD from an L1-lacZ reporter resulted in a loss of beta-galactosidase expression in the telencephalon and mesencephalon. Collectively, our experiments indicate that the HPD regulates L1 expression in neural tissues via homeodomain and Pax proteins and is likely to be a target of BMP signaling during development.  (+info)

Specification of distinct dopaminergic neural pathways: roles of the Eph family receptor EphB1 and ligand ephrin-B2. (6/1984)

Dopaminergic neurons in the substantia nigra and ventral tegmental area project to the caudate putamen and nucleus accumbens/olfactory tubercle, respectively, constituting mesostriatal and mesolimbic pathways. The molecular signals that confer target specificity of different dopaminergic neurons are not known. We now report that EphB1 and ephrin-B2, a receptor and ligand of the Eph family, are candidate guidance molecules for the development of these distinct pathways. EphB1 and ephrin-B2 are expressed in complementary patterns in the midbrain dopaminergic neurons and their targets, and the ligand specifically inhibits the growth of neurites and induces the cell loss of substantia nigra, but not ventral tegmental, dopaminergic neurons. These studies suggest that the ligand-receptor pair may contribute to the establishment of distinct neural pathways by selectively inhibiting the neurite outgrowth and cell survival of mistargeted neurons. In addition, we show that ephrin-B2 expression is upregulated by cocaine and amphetamine in adult mice, suggesting that ephrin-B2/EphB1 interaction may play a role in drug-induced plasticity in adults as well.  (+info)

A glial cell line-derived neurotrophic factor-secreting clone of the Schwann cell line SCTM41 enhances survival and fiber outgrowth from embryonic nigral neurons grafted to the striatum and to the lesioned substantia nigra. (7/1984)

We have developed a novel Schwann cell line, SCTM41, derived from postnatal sciatic nerve cultures and have stably transfected a clone with a rat glial cell line-derived neurotrophic factor (GDNF) construct. Coculture with this GDNF-secreting clone enhances in vitro survival and fiber growth of embryonic dopaminergic neurons. In the rat unilateral 6-OHDA lesion model of Parkinson's disease, we have therefore made cografts of these cells with embryonic day 14 ventral mesencephalic grafts and assayed for effects on dopaminergic cell survival and process outgrowth. We show that cografts of GDNF-secreting Schwann cell lines improve the survival of intrastriatal embryonic dopaminergic neuronal grafts and improve neurite outgrowth into the host neuropil but have no additional effect on amphetamine-induced rotation. We next looked to see whether bridge grafts of GDNF-secreting SCTM41 cells would promote the growth of axons to their striatal targets from dopaminergic neurons implanted orthotopically into the 6-OHDA-lesioned substantia nigra. We show that such bridge grafts increase the survival of implanted embryonic dopaminergic neurons and promote the growth of axons through the grafts to the striatum.  (+info)

Xenopus axin interacts with glycogen synthase kinase-3 beta and is expressed in the anterior midbrain. (8/1984)

Axin is encoded by the fused locus in mice and is required for normal vertebrate axis formation. It has recently been shown that axin associates with APC, beta-catenin and glycogen synthase kinase-3 (GSK-3) in a complex that appears to regulate the level of cytoplasmic beta-catenin. We have identified the Xenopus homologue of axin through its interaction with GSK-3b. Xenopus axin (Xaxin) is expressed maternally and throughout early development with a low level of ubiquitous expression. Xaxin also shows remarkably high expression in the anterior mesencephalon adjacent to the forebrain-midbrain boundary.  (+info)