Subcellular localization of full-length and truncated Trk receptor isoforms in polarized neurons and epithelial cells.
(17/1624)
Neurotrophins affect neuronal development and plasticity via spatially localized effects, yet little is known about the subcellular distribution of the Trk neurotrophin receptors and the impact of this distribution on neurotrophin action. To address this, we examined the subcellular location of full-length TrkB and TrkC tyrosine kinase receptors and truncated TrkB isoforms after transfection of Madin-Darby canine kidney (MDCK) cells, dissociated primary hippocampal neurons, and cortical neurons within intact brain slices. Myc-, herpes virus glycoprotein (HVG)-, or FLAG-derived epitope-tagged receptor isoforms were created to allow their unambiguous identification and localization after transfection. All tagged receptors were appropriately synthesized, and full-length myc-TrkB and myc-TrkC mediated appropriate neurotrophin-signaling events. We found that full-length TrkB receptors were excluded from the apical domain of MDCK cells but that TrkC receptors were present in both apical and basolateral domains. Full-length TrkB and TrkC were found throughout transfected primary cultured hippocampal neurons and transfected neurons in neocortical brain slices and showed no evidence of vectorial sorting. Truncated forms of TrkB were also homogeneously distributed in MDCK cells, dissociated hippocampal neurons, and cortical neurons within slice preparations. Levels of full-length and truncated TrkB were examined in postsynaptic densities; both receptor isoforms were present but only moderately enriched in these structures. Together, these findings suggest that Trk receptors are uniformly distributed in both axonal and dendritic compartments and that local neurotrophin responses are controlled by other mechanisms. (+info)
Sarcoplasmic reticulum Ca(2+) release by 4-chloro-m-cresol (4-CmC) in intact and chemically skinned ferret cardiac ventricular fibers.
(18/1624)
The purpose of this study was to determine whether 4-chloro-m-cresol (4-CmC) could generate caffeine-like responses in ferret cardiac muscle. The concentration dependence of 4-CmC-mediated release of Ca(2+) from the sarcoplasmic reticulum was studied in intact cardiac trabeculae and saponin-skinned fibers in which the sarcoplasmic reticulum was loaded with Ca(2+). In intact and saponin-skinned preparations isolated from right ventricle, the effect of 4-CmC on sarcoplasmic reticulum Ca(2+) content was estimated by analysis of caffeine contracture after application of chlorocresol. In addition, the effects of 4-CmC on maximal Ca(2+)-activated tension and the Ca(2+) sensitivity of myofibrils were analyzed by using Triton-skinned cardiac fibers. The results show that 4-CmC generates a contractile response in saponin-skinned but not intact fibers. The sarcoplasmic reticulum is implicated in the 4-CmC response; more precisely, in Ca(2+) release via the ryanodine receptor. Moreover, 4-CmC, like caffeine, has effects on maximal Ca(2+)-activated tension and the Ca(2+) sensitivity of myofibrils. (+info)
Interspecies differences in the cardiac negative inotropic effects of beta(3)-adrenoceptor agonists.
(19/1624)
The aim of the present study was to compare the effects of three preferential (BRL 37344, SR 58611, CL 316 243) and a partial (CGP 12177) beta-adrenoceptor (beta(3)-AR) agonists on the contractility of ventricular strips sampled from various mammalian species including humans. In the human heart, all beta(3)-AR agonists tested decreased contractility by 40 to 60% below control with an order of potency: BRL 37344 > CL 316 243 = SR 58611 >> CGP 12177. In the dog, the negative inotropic effects produced by beta(3)-AR stimulation were less pronounced than in humans, approximately 30% below control. The order of potency of beta(3)-AR agonists was CGP 12177 > BRL 37344 = SR 58611 >> CL 316 243; i.e., very different from that observed in humans. In rat, only BRL 37344 was efficient to decrease contractility. In guinea pig, only CL 316 243 significantly reduced peak tension. In both species, the reduction in peak tension did not exceed 20 to 30%. Finally, in the ferret, none of the agonists tested induced a negative inotropic effect. In dog, the negative inotropic effects of CGP 12177 were not modified by nadolol, but were abolished by bupranolol, a beta(1-3)-AR. beta(3)-AR transcripts were detected in the dog but not in the rat ventricle by using a reverse transcription-polymerase chain reaction assay. We conclude that cardiac negative inotropic effects related to beta(3)-AR agonist stimulation vary markedly depending on the species. A comparable interspecies variation previously has been reported concerning the lipolytic effects of beta(3)-AR agonist stimulation. Our study demonstrates that the pharmacological profile of a beta(3)-AR agonist on the human myocardium cannot be extrapolated from usual animal models. (+info)
Cross-modal reorganization of callosal connectivity without altering thalamocortical projections.
(20/1624)
Mammalian cerebral cortex is composed of a multitude of different areas that are each specialized for a unique purpose. It is unclear whether the activity pattern and modality of sensory inputs to cortex play an important role in the development of cortical regionalization. The modality of sensory inputs to cerebral cortex can be altered experimentally. Neonatal diversion of retinal axons to the auditory thalamus (cross-modal rewiring) results in a primary auditory cortex (AI) that resembles the primary visual cortex in its visual response properties and topography. Functional reorganization could occur because the visual inputs use existing circuitry in AI, or because the early visual inputs promote changes in AI's circuitry that make it capable of constructing visual receptive field properties. The present study begins to distinguish between these possibilities by exploring whether the callosal connectivity of AI is altered by early visual experience. Here we show that early visual inputs to auditory thalamus can reorganize callosal connections in auditory cortex, causing both a reduction in their extent and a reorganization of the pattern. This result is distinctly different from that in deafened animals, which have widespread callosal connections, as in early postnatal development. Thus, profound changes in cortical circuitry can result simply from a change in the modality of afferent input. Similar changes may underlie cortical compensatory processes in deaf and blind humans. (+info)
Correlational structure of spontaneous neuronal activity in the developing lateral geniculate nucleus in vivo.
(21/1624)
The properties of spontaneous activity in the developing visual pathway beyond the retina are unknown. Multielectrode recordings in the lateral geniculate nucleus (LGN) of awake behaving ferrets, before eye opening, revealed patterns of spontaneous activity that reflect a reshaping of retinal drive within higher visual stages. Significant binocular correlations were present only when cortico-thalamic feedback was intact. In the absence of retinal drive, cortico-thalamic feedback was required to sustain correlated LGN bursting. Activity originating from the contralateral eye drove thalamic activity far more strongly than that originating from the ipsilateral eye. Thus, in vivo patterns of LGN spontaneous activity emerge from interactions between retina, thalamus, and cortex. (+info)
Stimulation of Na+-K+-2Cl- cotransport by arsenite in ferret erythrocytes.
(22/1624)
1. Na+-K+-2Cl- cotransport activity was measured in ferret erythrocytes as the bumetanide-sensitive uptake of 86Rb. 2. The Na+-K+-2Cl- cotransport rate was stimulated by treating erythrocytes with sodium arsenite but not by sodium arsenate (up to 1 mM). Stimulation took an hour to develop fully. Arsenite had no effect on bumetanide-resistant 86Rb uptake. 3. In cells stored for 3 days or less, cotransport stimulation by arsenite could be described by assuming arsenite either acts at a single site (EC50, 60+/-14 microM, mean +/- S.E.M., n = 3) or that it acts at both high- (EC50, 35+/-9 microM, mean +/- S.E.M., n = 3) and low- (EC50 >2 mM) affinity sites. 4. Stimulation by 1 mM arsenite was greatest on the day of cell collection (rate about 3 times that of the control), even exceeding that produced by 20 nM calyculin A, and declined during cell storage. Addition of calyculin A to arsenite-stimulated cells resulted in further stimulation of Na+-K+-2Cl- cotransport, suggesting that arsenite and calyculin act synergistically. This was most apparent in stored cells. 5. Stimulation by 1 mM arsenite was not affected by treating cells with the mitogen-activated protein kinase inhibitors SB203580 (20 microM) and PD98059 (50 microM), but was both prevented and reversed by the kinase inhibitors staurosporine (2 microM), 4-amino-5-(4-methylphenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP1, 50 microM) and genistein (0.3 mM), and with a combination of 10 microM A23187 and 2 mM EDTA (to reduce intracellular Mg2+ concentration). Only treatment with EDTA and A23187 prevented stimulation by the combination of 1 mM arsenite and 20 nM calyculin, whereas no treatment was able to fully reverse this stimulation once elicited. 6. Our data are consistent with arsenite stimulating (perhaps indirectly) a kinase that phosphorylates and activates the Na+-K+-2Cl- cotransporter. (+info)
The critical period for ocular dominance plasticity in the Ferret's visual cortex.
(23/1624)
Microelectrode recordings and optical imaging of intrinsic signals were used to define the critical period for susceptibility to monocular deprivation (MD) in the primary visual cortex of the ferret. Ferrets were monocularly deprived for 2, 7 or >14 d, beginning between postnatal day 19 (P19) and P110. The responses of visual cortical neurons to stimulation of the two eyes were used to gauge the onset, peak, and decline of the critical period. MDs ending before P32 produced little or no loss of response to the deprived eye. MDs of 7 d or more beginning around P42 produced the greatest effects. A rapid decline in cortical susceptibility to MD was observed after the seventh week of life, such that MDs beginning between P50 and P65 were approximately half as effective as those beginning on P42; MDs beginning after P100 did not reduce the response to the deprived eye below that to the nondeprived eye. At all ages, 2 d deprivations were 55-85% as effective as 7 d of MD. Maps of intrinsic optical responses from the deprived eye were weaker and less well tuned for orientation than those from the nondeprived eye, with the weakest maps seen in the hemisphere ipsilateral to the deprived eye. Analysis of the effects of 7 d and longer deprivations revealed a second period of plasticity in cortical responses in which MD induced an effect like that of strabismus. After P70, MD caused a marked loss of binocular responses with little or no overall loss of response to the deprived eye. The critical period measured here is compared to other features of development in ferret and cat. (+info)
Maps of central visual space in ferret V1 and V2 lack matching inputs from the two eyes.
(24/1624)
In the visual cortex, the representation of central visual space is supplied by matching geniculate inputs that are driven exclusively by one eye or the other. In layer 4 of early visual areas (V1 in primates and V1 and V2 in cat), these inputs form a nearly uniform array of small ocular dominance domains, while preserving overall topographic order within the cortical map. In ferret, however, ocular dominance domains in different regions of the visual cortex are strikingly irregular in size and shape. The exceptionally large size of domains in some regions implies a departure from the usual visuotopic matching of inputs from the two eyes. Using optical-imaging, electrophysiological, and anatomical techniques, we show that this regional variation is attributable to exclusively monocular maps of the central portions of the ipsilateral visual field in V1 and the contralateral visual field in V2. In addition, we document a complex interdigitation of V1 and V2 that entails a discontinuity in the mapping of visual space and fragmentation of V2 into isolated cortical territories. We suggest that both the monocularity of these cortical maps and the visuotopic discontinuity along the V1-V2 border derive from asymmetries in the crossed and uncrossed retinal pathways. (+info)