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T Gotow

Publications and source records attributed to T Gotow.

At least 19 recordsLinked to original sources
PubMed1995

The organization of neurofilaments accumulated in perikaryon following aluminum administration: relationship between structure and phosphorylation of neurofilaments.

Neurofilaments accumulated in perikarya and dendrites of anterior horn cells and Purkinje cells of rabbit treated by aluminum chloride were analysed with a variety of techniques. Four different monoclonal antibodies against phosphorylated and nonphosphorylated epitopes on neurofilament H subunit were used to compare phosphorylation state of these accumulated neurofilaments with that of axonal neurofilaments. Although immunoblotting revealed no significant difference in phosphorylation between control and aluminum-treated brains, accumulated neurofilaments were immunocytochemically more phosphorylated than control perikaryal or dendritic neurofilaments. With detailed analysis of cryothin-section immunogold labeling, accumulated neurofilaments were, however, significantly less phosphorylated than axonal neurofilaments. With quick-freeze deep etching, core filaments of accumulated neurofilaments are as dense as axonal neurofilaments but much less regularly aligned. Cross-bridges of accumulated neurofilaments were less frequent and more branched than those of axonal neurofilaments, and when examined with combined immunocytochemistry and deep etching, were less phosphorylated. These results suggest that there is a relationship between the phosphorylation and the structural organization of neurofilaments. The phosphorylation of neurofilament H subunit may be necessary for formation of frequent and straight cross-bridges and resulting regular alignment of core filaments.

Aluminum↗
PubMed1994

Single K+ channels closed by light and opened by cyclic GMP in molluscan extra-ocular photoreceptor cells.

We report the first recordings of the light-sensitive channel which is active during dark and is closed by light in the Onchidium extra-ocular photoreceptor cells. This light-sensitive channel was K-selective and was not blocked by extracellular Ca2+ and Mg2+. Application of cyclic GMP to excised inside-out patches activated (opened) a channel that appeared to be the same as the light-sensitive channel recorded from the same membrane in the intact cell.

Animals↗
PubMed1994

Mechanisms of hyperpolarization induced by two cytokines, hTNF alpha and hIL-1 alpha in neurons of the mollusc, Onchidium.

The voltage and current responses induced by extracellular tumor necrosis factor (hTNF alpha) or interleukin-1 (hIL-1 alpha) on the Be-1 and Es-1 neurons of the Onchidium ganglia were examined. Pressure-ejected hTNF alpha or hIL-1 alpha produced an inhibitory, hyperpolarized effect in unclamped neurons. In the same neurons voltage-clamped at their resting potential levels, the same hTNF alpha or hIL-1 alpha elicited an outward current having a time course similar to that of the hyperpolarization, associated with a decreased membrane conductance. The hTNF alpha- or hIL-1 alpha-induced outward current did not reverse even at positive membrane potentials considerably above + 100 mV in the absence ouabain (a specific blocker of Na-pump). In the presence of ouabain, the hTNF alpha- or hIL-1 alpha-induced current was reduced over a wide range of membrane potential, so that the current reversed at about + 20 mV. Lowering the external Na+ concentration from 450 to 200 mM in the presence of ouabain, shifted the reversal potential from + 20 to 0 mV, to near the shift value of 20.8 mV predicted by the Nernst equation for a Na(+)-selective conductance. Neither an increase nor a decrease of extracellular K+, Cl- or Ca2+, however, significantly altered the current induced by hTNF alpha or hIL-1 alpha. These suggest that the hTNF alpha- or hIL-1 alpha-induced hyperpolarization or outward current response is mediated by two mechanisms, a decrease in Na+ conductance and activation of the Na-pump.

Animals↗
PubMed1994

Phosphorylation of neurofilament H subunit as related to arrangement of neurofilaments.

To find out what causes differences in phosphorylation states in neurofilaments (NF), we selected two types of dendrite, one provided with very few NFs (Purkinje cell) and the other with relatively many (anterior horn cell). We examined these with four monoclonal antibodies selected by the Western blot analysis, two (NE14 and SMI31) recognizing only phosphorylated, SMI32 recognizing only nonphosphorylated, and N52 recognizing phosphorylation-independent epitopes of NF-H. The immunoperoxidase labeling of dendrites, and also of perikarya, in both neurons was detectable with all four antibodies. After the tissue was treated with Triton X-100, the labeling was still detectable with SMI32 or N52, but undetectable with NE14 and SMI31. The brain homogenate Triton-extracted supernatant after centrifugation at 100,000g for 1 hr showed the staining of NE14, SMI31, and N52 but not that of SMI32. In Purkinje cell dendrite and perikaryon, NFs always appeared singly. In the immunogold labeling, they were labeled only with SMI32 or N52. Labeling by NE14 or SMI31 was distributed throughout the cytoplasm and hardly associated with NFs. In the anterior horn cell dendrite and perikaryon, NFs appeared both singly and in bundles. They were predominantly labeled with SMI32 or N52 when they were single, and with NE14, SMI31, or N52 when they were bundled. Even in one NF, portions that appeared single were labeled mostly with SMI32 or N52, while the remainder, to which other NFs approached closely, were labeled mostly with NE14, SMI31, or N52. Thus, when NFs appear singly, NF-H in their projections or cross-bridges with other organelles is not phosphorylated, while when NFs are bundled, NF-H is phosphorylated in crossbridges between NF core filaments. These data may explain why the NF-H is heavily phosphorylated in axons, where NFs are abundant, and not in dendrites and perikarya, where NFs are sparse.

Animals↗
PubMed1994

Electrical myotonia of rabbit skeletal muscles by HMG-CoA reductase inhibitors.

HMG-CoA reductase (HCR) inhibitors are effective cholesterol-lowering agents in the treatment of hypercholesterolemia. Using intracellular microelectrodes, we studied the pathomechanism of myotonia experimentally induced in rabbits by HCR inhibitors, simvastatin, and pravastatin. The external intercostal muscle of rabbits showed some electrophysiologic characteristics of myotonia including repetitive firing after administration of simvastatin (50 mg/kg per day, for 4 weeks). The relative chloride conductance, though reduced in both, was more affected in simvastatin-administered muscles. In normal muscles perfused with a solution containing the inhibitors, both simvastatin and pravastatin produced membrane hyperexcitability with repetitive firing similar to that seen in simvastatin-administered rabbits. The minimum concentrations required to cause repetitive firing was 0.3 mg/L for simvastatin and 30 mg/L for pravastatin. These results indicate that HCR inhibitors induce some characteristics of myotonia by blocking the chloride channel in the muscle membrane.

Action Potentials↗
PubMed1994

Dephosphorylation of the largest neurofilament subunit protein influences the structure of crossbridges in reassembled neurofilaments.

Phosphorylation-dependent change in electrophoretic mobility is the most unique characteristic of NF-H, the largest molecular mass subunit of the neurofilament. We dephosphorylated NF-H using Escherichia coli alkaline phosphatase, then reassembled it into neurofilaments with NF-M and NF-L, and into NF-H filaments with NF-H alone. We compared these dephosphorylated filaments with control: projections by low-angle rotary-shadow, crossbridges by quick-freeze deep-etch, and core filament packing density by thin-section electron microscopy. Projections in the dephosphorylated filaments were basically similar in structure to those in control, although there was a tendency for them to be wider and less dense, especially in NF-H filaments. Dephosphorylated filaments were still able to form crossbridges between core filaments, but their crossbridges were significantly wider, less dense, more branched and more irregular than crossbridges in control, and core filaments were more densely packed. These structural differences may be brought about by the removal of phosphate groups from NF-H tail and consequent reduction of electrostatic repulsion between adjacent crossbridges extending from the same core filament. The results indicate that phosphorylation of NF-H is necessary for forming well developed crossbridges, straight and at constant intervals, like those of in vivo axonal neurofilaments.

Actin Cytoskeleton↗
PubMed1992

Macromolecular structure of reassembled neurofilaments as revealed by the quick-freeze deep-etch mica method: difference between NF-M and NF-H subunits in their ability to form cross-bridges.

Neurofilament (NF) structure and ability to form cross-bridges were examined by quick-freeze deep-etch mica and low-angle rotary-shadow electron microscopy in NFs purified from bovine spinal cord and reassembled in various combinations of NF subunits. When NFs were reassembled from triplet proteins, NF-L, NF-M and NF-H, they were oriented randomly and often fragmented, but their elongated filaments (12-15 nm wide) and the cross-bridges (4-5 nm wide) connecting them were similar in appearance to those of isolated bovine NFs or in vivo rat NFs. Projections extended from the wall of the core filament in almost the same pattern as the cross-bridges and were the same in width and interval (minimum interval, 20-25 nm) as the cross-bridges. Projections were more conspicuous when core filaments were separated by 60 to 80 nm or more, while cross-bridges were more conspicuous when core filaments were close to each other. Projections or cross-bridges extended bilaterally at intervals of 20 to 25 nm where core filaments expanded and formed a network between filaments which were far from one another. When NFs were reconstructed from NF-L alone, only core filaments appeared, the same width as the filaments of triplet NFs. The core filaments were occasionally in almost direct contact with each other, with no projection or cross-bridge. When NFs were reassembled from NF-M alone or NF-L + NF-M, although NF-M core filaments were shorter and slightly thinner than NF-L + NF-M core filaments, both had projections, and both had cross-bridges, but cross-bridges were less evident. Cross-bridges were almost the same in width as those of triplet NFs, but significantly shorter and much less frequent although the minimum interval was the same, and core filaments were not attached to each other. In contrast, when NFs were reconstituted from NF-H alone or NF-L + NF-H, both had conspicuous projections and cross-bridges, similar to those of triplet NFs. Thus, when NFs contained NF-H, they formed frequent cross-bridges and long projections with extensive peripheral branching. When NFs contained NF-M but no NF-H, they tended to form cross-bridges, and to form projections that were shorter and straighter and without peripheral branching. That is, there appears to be a significant difference between NF-M and NF-H in ability to form cross-bridges and thus in interaction with adjacent NFs.

Animals↗
PubMed1991

Roles of cyclic GMP and inositol trisphosphate in phototransduction of the molluscan extraocular photoreceptor.

The internal messengers mediating the photocurrent of the molluscan extraocular photoreceptor, A-P-1, were examined. In the dark, pressure-injection of cGMP into the A-P-1, voltage-clamped at resting levels, produced a rapid outward current, associated with an increase in conductance. However, the cGMP-induced current and increase in conductance were suppressed by subsequent photostimulation, suggesting hydrolysis of cGMP by light. The steady-state I/V relation for the cGMP-induced current was non-linear. The I/V relation for the instantaneous cGMP-induced current, measured 50 ms after the beginning of a voltage step, was linear, and reversed at the membrane potential, -67 mV, which corresponded to the K+ equilibrium potential of A-P-1 in 10 mM K+ normal saline. These findings indicate that the internal cGMP induces a voltage- and time-dependent K+ current. Since the photocurrent results from the suppression of a voltage- and time-dependent K+ current similar to above, the photocurrent is considered to be equivalent to the suppression of the cGMP-induced current. Short pressure-injection of GDP-beta-S into A-P-1 reduced the subsequent photocurrent. The photocurrent was also suppressed after an external application of Pertussis toxin. On the other hand, the photocurrent was amplified by prior pressure-injection of inositol 1,4,5-trisphosphate (IP3). However, a short pressure-injection of neomycin into A-P-1 depressed the subsequent photocurrent. These results suggested that the cGMP-induced (dark) current is mediated by cGMP, and that hydrolysis of cGMP by light leads to the photocurrent, then being modified by another messenger, IP3, to be amplified.(ABSTRACT TRUNCATED AT 250 WORDS)

1-Methyl-3-isobutylxanthine↗
PubMed1991

Cytoplasmic architecture of the axon terminal: filamentous strands specifically associated with synaptic vesicles.

Cytoplasmic architecture of axon terminals in rat central nervous tissue was examined by quick-freeze deep-etch method to determine how synaptic vesicles and their associated cytoplasmic environment are organized in the terminal and to know how these structures participate in the mechanism for neurotransmitter release. The axoplasm is divisible into two domains: one occupied by mitochondria in the middle of the terminal, called the mitochondrial domain, the other situated in the periphery and exclusively filled with spherical synaptic vesicles, 50-60 nm in diameter, the synaptic vesicle domain. The most characteristic feature of the mitochondrial domain was the appearance of many microtubules connected with mitochondria by filamentous strands. Large vesicles, 80-100 nm in diameter, were preferentially associated with the mitochondrial domain, and linked with microtubules wherever they appeared. The cytoplasmic matrix of the synaptic vesicle domain showed a more fibrillar texture than that of the mitochondrial domain because of the distribution of filamentous strands associated with synaptic vesicles. These strands were significantly thicker and longer (mean 11.7 nm thick and 42.7 nm long) than those linking membrane-bound organelles to microtubules (mean 8.3 nm thick and 23.0 nm long), and connected vesicles to one another or to the plasma membrane, making a complicated network around the vesicles. Further, both strands were significantly different in dimension from actin filaments (mean 9.9 nm thick and 73.5 nm long) showing 5-nm axial periodicity. These strands, especially synaptic vesicle-associated ones including their network, were readily broken down in the most part by detergent treatment or chemical fixation, indicating that they are very delicate in nature. Granular materials, which are spherical and vary in size (6-20 nm in diameter), are also more conspicuous in the synaptic vesicle domain than in the mitochondrial domain. More fibrillar and granular cytoplasmic structure of the synaptic vesicle domain may be crucial for synaptic vesicles to perform an essential role in releasing the transmitter.

Animals↗
PubMed1991

Characterization of intracytoplasmic neurofilament accumulation in hamster brain caused by Alzheimer buffy coat inoculation: comparison with experimental neurofibrillary changes produced by aluminum intoxication.

Neurofilament accumulation was produced in neuronal perikarya of the lower brain stem nuclei in the hamster brain by inoculating buffy coat from a patient with Alzheimer's disease. The abnormally proliferated neurofilament was characterized by immunostaining and electron microscopy. The intracytoplasmic neurofilament accumulation was positively stained with anti-tau and anti-ubiquitin, but not with antibody against paired helical filaments. Experimental neurofibrillary changes were produced in the rabbit brain by aluminum injection, and the proliferated neurofilaments in the rabbit brain were compared with those in the hamster brain inoculated with Alzheimer buffy coat. The results indicate that the long pathological process in the hamster brain is triggered by Alzheimer's buffy coat inoculation which results in intracytoplasmic accumulation of modified neurofilaments.

Aluminum↗
PubMed1990

Differential distribution of serotoninergic inputs on the goldfish Mauthner cell.

The morphology and distribution of the serotoninergic (5-HT) input to the Mauthner cell (M cell) of a teleost, Carassius auratus, were analyzed at the light microscopic level. Immunohistochemical methods revealed that 1) most fibers innervating the M cell originate from the ventral and lateroventral regions of the rhombencephalon; 2) two groups of fibers contribute to this innervation, thick ones (type I, 0.4-0.7 microns in diameter) with terminal endings and thin ones (type II, less than 0.2 microns) that issue numerous beaded varicosities 4-10 microns from the target cell and only occasional side endings contacting it; 3) the density of immunoreactive profiles is uneven over the whole cell and predominates on the ventral dendrite; and 4) the two sets of axons, although overlapping, do not have the same distribution. Specifically, both classes are present on the ventral dendrite, whereas type II fibers are the only ones observed on the soma, in the region of the initial segment of the axon, and in the vicinity of the lateral dendrite. Functionally identified inputs on the M cell also have a regionalized distribution, depending, for example, on whether they belong to excitatory or inhibitory networks. Thus we propose that 5-HT inputs have specific influences that are a function of their respective localization.

Animals↗
PubMed1990

The light-suppressible K+ conductance and evaluation of internal messenger candidates in the molluscan extraocular photoreceptor.

A photoreceptor potential produced by a decrease in membrane conductance was not thought to occur in any invertebrate photoreceptors. However, we have found that the molluscan extraocular photoreceptor, A-P-1 responds to light with a depolarizing receptor potential due to a decrease in K+ conductance, so that the photoresponse associated with a decrease in membrane conductance is not unique to the vertebrate photoreceptor. The properties that the light-suppressible K+ conductance is time- and voltage-dependent are explained by comparison with those of the single channel conductance obtained in patch-clamp of both vertebrate and invertebrate photoreceptors. The noise analysis of the light-induced current suggest that this macroscopic light-suppressible conductance consists of channels. It is concluded that the light-suppressible K+ conductance is mediated by hydrolysis of cGMP which reduces internal cGMP, in agreement with the cGMP hypothesis of vertebrate phototransduction and that the hydrolysis may be modified by IP3.

Animals↗
PubMed1989

Collateral sprouting of somatostatin-immunoreactive axons after partial deafferentation of the central nucleus of the rat amygdala.

These experiments utilize a paradigm developed to study plastic responses of peptidergic neurons in a discrete brain area following deafferentation. The central nucleus of the amygdala (CNA) is richly innervated by somatostatin-immunoreactive (SS-I) terminal axons. In the course of preliminary light microscopic (LM) investigations by this laboratory, changes were observed in the density of presumed SS-I terminals in the rat CNA after lesioning the medial input. The LM finding of increased density of presumed SS-I terminals in the CNA at the 10-day post-lesion stage underscored the need for a quantitative electron microscopic (EM) study of the SS-I components, including an evaluation of synaptic events at different survival periods. At the 3-day post-lesion stage, EM examination showed degenerating axons in the lesioned CNA, many already engulfed by astrocytes. None of the degenerating profiles were SS-I, supporting the view that the lesion did not interrupt, to any significant extent, SS-I axons entering the nucleus. EM surveys of the 10-day post-lesion material demonstrated that degenerated profiles had almost completely disappeared. Numbers of SS-I axon terminals, particularly of smaller-sized profiles, were increased by 22% over control value. Synaptic frequency was decreased by 16% below control value. Numbers of SS-I terminals making synapses were increased 3.4% above control value. At the 30-day post-lesion stage, the total number of SS-I terminal axons had increased 86% over controls, whereas the synaptic frequency had decreased by about a third below controls. The absolute number of SS-I terminals engaging in synapses had increased by 24% over controls. The 90-day post-lesion CNA showed a further increase in the number of SS-I axon profiles: 136% over control value. The synapse-to-axon ratio (synaptic frequency) of 27% was similar to that observed for the CNA from the unlesioned side or from unoperated animals. At this stage the number of SS-I synapses had increased by 135% over controls. This model presents many possibilities for studying neuroplasticity, particularly involving peptidergic neurons of the central autonomic nervous system.

Amygdala↗
PubMed1989

Effect of serotonergic afferents on quantal release at central inhibitory synapses.

Although most examples of modulation of synaptic transmission have been obtained from excitatory rather than from inhibitory connections, serotonin (5HT) is now shown to cause a presynaptic facilitation of release of the inhibitory neurotransmitter glycine. Brief local injections of this amine, or application of a 5HT uptake blocker, produce a long-lasting enhancement of both spontaneous and evoked inhibitory currents in the teleost Mauthner cell. Quantal analysis showed that the probability of release is increased. Focal recording indicated that 5HT acts directly on the inhibitory terminals, possibly reducing potassium conductances. Double staining with specific antibodies demonstrated a morphological substrate for this effect. Nerve endings that contain 5HT contact inhibitory terminals directly apposed to postsynaptic glycine receptors.

Afferent Pathways↗
PubMed1989

A light-induced decrease of cyclic GMP is involved in the photoresponse of molluscan extraocular photoreceptors.

The depolarizing photoreceptor potential in the molluscan extraocular photoreceptor, A-P-1, results from the light-induced suppression of specific K+ currents. An application of cGMP or IBMX (inhibitor of phosphodiesterase) to A-P-1-evoked light-suppressed currents, similar to the above specific K+ currents. Thus, the light-induced decrease of cGMP levels in A-P-1 may be responsible for the photoreceptor potential, like vertebrate phototransductions.

1-Methyl-3-isobutylxanthine↗
PubMed1989

Photoresponses of an extraocular photoreceptor associated with a decrease in membrane conductance in an opisthobranch mollusc.

The photoresponse of an extraocular photoreceptor, the photoresponsive neuron (A-P-1) in the abdominal ganglion of Onchidium verruculatum, was studied by using a voltage-clamp with two micropipettes and a monochromatic light. When the A-P-1 was voltage-clamped at resting membrane potential levels, light induced a slowly developing inward current which peaked at about 20 s. A decrease in membrane conductance accompanied this light-induced current which corresponded to the depolarizing photoreceptor potential in the unclamped A-P-1. The relationship between the peak of the current response and light intensity could be predicted by using the modified Michaelis-Menten equation. The spectral sensitivity for the photoresponse had a peak at 490 nm. The steady-state light-induced current was a non-linear function of the membrane potential. The current-voltage relationship for the instantaneous light-induced current was almost linear. In normal (10 mM K+) saline, the polarity of the instantaneous light current reversed from inward to outward at about -67 mV, and doubling the external K+ from 10 to 20 mM shifted the reversal potential to about-50 mV, similar to that predicted by a K+-electrode. These results suggest that the light-induced current or the depolarizing receptor potential of A-P-1 is due to the light suppression of a voltage- and time-dependent K+ current.

Animals↗
PubMed1989

Orthogonal arrays of particles in plasma membranes of Müller cells in the guinea pig retina.

Plasma membranes of guinea pig Müller cells were examined with a freeze-fracture technique to see how orthogonal arrays are distributed in the avascular retina. Examination of the portion approximately intermediate between the optic disc and equator of the eyeball showed that all end-feet of Müller cells were provided with arrays. Orthogonal arrays were concentrated on vitreal end-foot membranes, i.e., membranes that were covered by the basal lamina and contacted the vitreous body, called vitreal membranes here. The arrays were rarely observed in the portions of end-feet that did not contact the vitreous body, called lateral membranes. The distribution density of arrays in the vitreal membranes was 122.5 +/- 45.3/microns2, which was over 10 times higher than that (9.6 +/- 9.6/microns2) in the lateral membranes. The arrays became numerous and extended in shape at the periphery of the vitreal membrane, characteristically aligned in rows at the border where vitreal met lateral membrane, but never intruded into the domain of lateral membrane. Some arrays were composed of loosely attached particles and/or rod-like profiles. Sometimes rod-like profiles, 9-13 nm wide and 20-50 nm long, called linear structures here, were isolated, and sometimes they appeared in rows. Ordinary intramembrane particles (IMPs) were significantly smaller and less numerous in vitreal than in lateral membranes. IMPs larger than 9 nm in diameter were significantly fewer in the vitreal membranes, which suggests that they have been consumed to form the arrays. Although the distribution of orthogonal arrays is similar to that of K+ channels (Newman: J. Neurosci., 7:2423-2432, 1987), we consider the array an unlikely candidate for the ion channel, because its subunit particles do not protrude onto either the inner or outer surface of the membrane (Gotow and Hashimoto: J. Neurocytol., 17:399-413, 1988). Judging from their unique alignment in rows where the membrane is bent and vitreal and lateral membranes meet, the arrays may contribute to some membrane stability, resisting the physical tension at the interface with mesenchymal tissue.

Animals↗