Neurotrophins and their Trk‐receptors in the cerebellum of zebrafish

Neurotrophins (NTs) and their specific Trk‐receptors are key molecules involved in the regulation of survival, proliferation, and differentiation of central nervous system during development and adulthood in vertebrates. In the present survey, we studied the expression and localization of neurotrophins and their Trk‐receptors in the cerebellum of teleost fish Danio rerio (zebrafish). Teleostean cerebellum is composed of a valvula, body and vestibulolateral lobe. Valvula and body show the same three‐layer structure as cerebellar cortex in mammals. The expression of NTs and Trk‐receptors in the whole brain of zebrafish has been studied by Western blotting analysis. By immunohistochemistry, the localization of NTs has been observed mainly in Purkinje cells; TrkA and TrkB‐receptors in cells and fibers of granular and molecular layers. TrkC was faintly detected. The occurrence of NTs and Trk‐receptors suggests that they could have a synergistic action in the cerebellum of zebrafish. J. Morphol. 277:725–736, 2016. © 2016 Wiley Periodicals, Inc.     

Neurotrophins are not required for normal embryonic development of olfactory neurons

Neurons of the vertebrate olfactory epithelium (OE) regenerate continuously throughout life. The capacity of these neurons to regenerate and make new and precise synaptic connections in the olfactory bulb provides a useful model to study factors that may control or mediate neuronal regeneration. Expression and in vitro studies have suggested potential roles for the neurotrophins in the olfactory system. To directly examine whether neurotrophins are required for olfactory neuron development, we characterized in vivo the role of the neurotrophins in the primary olfactory system. For this, we generated mutant mice for TrkA, TrkB, TrkC, and also for BDNF and NT3 together with P2-IRES-tau-LacZ trangenic mice. Histochemical staining for beta-galactosidase at birth allowed in vivo analysis of the P2 subpopulation of olfactory neurons as well as their projections to the olfactory bulb. Our data indicate that Trk signaling is not required for normal embryonic development of the olfactory system.     

Tectorial membrane changes in hypothyroid rats during postnatal development

The morphological changes of the tectorial membrane (TM) during the postnatal development (0, 3, 6, 12 and 25 day old) of the organ of Corti were studied by light microscopy in 20 control and hypothyroid rats. Hypothyroidism was induced by daily administration of propylthioruracil (PTU) until the end of lactation. The auditive receptor in the cochlea of the hypothyroid animals shows serious structural alterations compared with those of normal ones: abnormal persistence of K?lliker's organ, immaturity of sensory cells and supporting cells and a specific distortion of the TM. Differences with controls were first observed on the sixth postnatal day of the hypothyroid rats. The inner spiral sulcus was not shaped and the TM was attached to the K?lliker's organ. In older stages (12 and 25 days), K?lliker's organ was still present. The TM acquired a shap hump with an abnormal fibrillar arrangement in its middle part. It was still attached to the outer supporting cells by a remnant of the marginal net. It was suggested that the TM is secreted by the inner spiral limbus and K?lliker's organ. An abnormal persistence of these structures in the hypothyroidism results in a retardation of Corti's organ development. However, this conclusion does not explain the absence of the outer portion of the TM. Our study confirms the hypothesis that the secretion of any components of the marginal zone of TM is made by outer supporting cells which in PTU-treated animals appear very immature and with hypoplasia.     

Gelsolin immunoreactivity and development of the tectorial membrane in the cochlea of normal and hypothyroid rats

Summary Gelsolin was localized by immunocytochemistry in the developing cochlea of the rat. In normal animals, the protein appeared at 18 th day in utero in cells of the Kölliker's organ, which are involved in the secretion of the tectorial membrane. The Kölliker's organ cells were not immunoreactive after the first postnatal week, which is when they cease their secretory activity. Gelsolin immunoreactivity was similar in thyroid-deficient rats until the second postnatal week but, at this age, Kölliker's organ did not transform and its gelsolin immunoreactivity persisted, together with its secretory activity. As a result, the tectorial membrane was greatly distorted and out of contact with the hair cells, which dramatically impaired the mechanical properties of the organ of Corti. The developing cochlea thus provides an example of the involvement of gelsolin in a secretory process that is of importance in the development of hearing.     

Soman intoxication in hypothyroid rats: alterations in brain neuronal RNA,acetylcholinesterase and survival

Studies were conducted to investigate relationships among soman (pinacolyl methylphosphonofluoridate) induced seizure activity, central metabolic impairments and lethality in normal vs thyroid-deficient rats. Quantitative cytophotometric measurements of individual cerebrocortical (layer V) and striatal neuron RNA contents were made following dosages of 0.5, 0.9 and 1.5 LD₅₀ soman (LD₅₀ = 135 μg/kg, sc). Hypothyroidism was associated with a marked diminution of overt convulsive activity and reduced susceptibility to lethal actions of soman as indicated by enhanced 24- and 48-h survival rates at 0.9, 1.2 and 1.5 LD₅₀. Hypothyroidism per se produced RNA depletion in both cortical and striatal neurons. Soman treatment diminished cortical RNA to essentially the same extent in thyroid-deficient rats as in euthyroids, whereas there was no further reduction of striatal neuron RNA. It was found that amelioration of convulsive activity and lethal- ity in hypothyroid rats was accompanied by reduced cerebral acetylcholinesterase (AChE, EC 3.1.1.7) inactivation, and that plasma cholinesterase (EC 3.1.1.8) and aliesterase (EC 3.1.1.1) levels were significantly higher in hypothyroid than in euthyroid saline-control rats. The overall data indicate that soman- induced central metabolic impairments can occur independent of paroxysmal neural activity and lethal actions of the agent. Resistance to soman observed with thyroid deficiency may be due in large part to increased binding to plasma enzymes and diminished delivery of soman to AChE in vital cholinergic sites.     

Neurotrophins: the biological paradox of survival factors eliciting apoptosis

Neurotrophins are target-derived soluble polypeptides required for neuronal survival. Binding of neurotrophins to Trk receptor tyrosine kinases initiate signaling cascades that promote cell survival and differentiation. All family members bind to another receptor (p75NTR), which belongs to the tumor necrosis factor superfamily. Hence, nerve growth factor (NGF) and related trophic factors are unique in that two separate receptor types are utilized. Although the biological function of p75NTR has been elusive, it has been suggested to mediate apoptosis of developing neurons in the absence of Trk receptors. This presents a tantalizing paradigm, in which life-death decisions of cells are dependent upon the expression and action of two different receptors with distinctive signaling mechanisms. In the presence of TrkA receptors, p75 can participate in the formation of high affinity binding sites and enhanced NGF responsiveness leading to a survival signal. In the absence of TrkA receptors, p75 can generate, in only specific cell populations, a death signal. Here we discuss the unique features and implications of this unusual signal transduction system.     

Oligodendrocyte ablation affects the coordinated interaction between granule and Purkinje neurons during cerebellum development

Oligodendrocytes (OLs) are the glial cells of the central nervous system (CNS) classically known to be devoted to the formation of myelin sheaths around most axons of the vertebrate brain. We have addressed the role of these cells during cerebellar development, by ablating OLs in vivo. Previous analyses had indicated that OL ablation during the first six postnatal days results into a striking cerebellar phenotype, whose major features are a strong reduction of granule neurons and aberrant Purkinje cells development. These two cell types are highly interconnected during cerebellar development through the production of molecules that help their proliferation, differentiation and maintenance. In this article, we present data showing that OL ablation has major effects on the physiology of Purkinje (PC) and granule cells (GC). In particular, OL ablation results into a reduction of sonic hedgehog (Shh), Brain Derived Neurotrophic Factor (BDNF), and Reelin (Rln) expression. These results indicate that absence of OLs profoundly alters the normal cerebellar developmental program.     

Deiodase activity in hypothyroid rats]

In rats hypothyroidized with methylthiouracil (MTU), methimazol (MMI), or radiothyroidectomy, the extent of deiodination for L-diiodotyrosine (L-DIT) and L-thyroxine (L-T4) was investigated in homogenate supernatants of liver and kidney. Deiodination in liver and kidney for DIT is twice as high as for T4, but the kidney allows only 25% of the liver deiodination activity both for DIT and T4. In the livers of all hypothyroid animals, iodide splitting both from DIT and T4 is highly significantly reduced by one-half compared with controls. In the kidney of all hypothyroid animals, the DIT deiodination is highly significantly lowered in comparison with controls; the T4 deiodination is significantly reduced only in animals treated with MTU and MMI, and is not significantly enhanced in radiothyroidectomized rats. Thus, there is no difference between MTU and MMI in the extent of deiodination for DIT and T4 in the homogenate supernatants of rat liver or kidney.     

Taurine-activated currents in isolated neurons of the rat cerebellum

Taurine-activated currents were investigated in rat cerebellar neurons using techniques of voltage clamping at the membrane and intracellular perfusion. Activation of both chloride and calcium conductance at the membrane were produced by applying taurine to the membrane surface. The dose-response curve for taurine-activated current is in the 1×10^–4–1×10^–1 M concentration region. The dissociation constant of the taurine-receptor complex equals 2×10^–3 M. Activation of taurine-induced currents is a cooperative process: Hill's coefficient –2. It was found that bicuculline and strychnine exert a blocking action on taurine-activated currents, while pentobarbital and oxazepam potentiate taurine action.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 22, No. 6, November–December, 1990, pp. 780–786.     

Pituitary-thyroid function of fetuses of hypothyroid and growth hormone treated hypothyroid rats.

Maternal hypothyroidism induced by surgical thyroidectomy (Tx) of the rat resulted in significantly higher fetal serum levels of thyroid stimulating hormone (TSH) and thyroxine (T4) on day 22 of gestation. Surprisingly, administration of growth hormone (GH) to hypothyroid mothers increased further the fetal serum T4 and TSH. The in vitro uptake of 131I-T4 by erythrocytes was elevated significantly when incubated with serum from fetuses of both hypothyroid and hypothyroid GH-treated mothers. Although the plasma protein levels of hypothyroid mothers and their fetuses are decreased significantly as compared to controls this is not true of hypothyroid GH-treated mothers and their fetuses. The T4 levels of both groups of Tx mothers were significantly below that of controls. However, as in the case of their fetuses, the serum T4 of GH-treated hypothyroid mothers was elevated from that of Tx only animals. It is concluded that the pituitary-thyroid system of fetuses of hypothyroid mothers is activated excessively during late gestation, that considerable T4 can be transported from the fetus to the mother during this period and that these fetuses are in fact born in a hyperthyroid state which is aggravated by maternal treatment with GH.     

Basic fibroblast growth factor promotes the survival and development of mesencephalic neurons in culture

Neuronotrophic effects of basic fibroblast growth factor (bFGF) have been reported for some central nervous system neurons. Here we report that also rat mesencephalic neurons are responsive to bFGF. bFGF produces a significant increase in the number of neurite-bearing cells, as well as in the degree of their fiber network. The present findings also provide the first evidence that bFGF can affect, in a concentration-dependent manner, at least two defined CNS neuronal populations, i.e., dopaminergic and gabaergic neurons. This effect was quantified by assessing the specific [3H]dopamine and [gamma-14C]aminobutyric acid uptakes with time in culture. Stimulation of uptake was more pronounced for dopaminergic neurons, suggesting a relative specificity in the actions of bFGF. These effects of bFGF were completely blocked by affinity-purified polyclonal antibodies. The possibility that bFGF plays a key role in normal nervous system development or function is discussed.     

Glial cells maintain synaptic structure and function and promote development of the neuromuscular junction in vivo

To investigate the in vivo role of glial cells in synaptic function, maintenance, and development, we have developed an approach to selectively ablate perisynaptic Schwann cells (PSCs), the glial cells at the neuromuscular junction (NMJ), en masse from live frog muscles. In adults, following acute PSC ablation, synaptic structure and function were not altered. However, 1 week after PSC ablation, presynaptic function decreased by approximately half, while postsynaptic function was unchanged. Retraction of nerve terminals increased over 10-fold at PSC-ablated NMJs. Furthermore, nerve-evoked muscle twitch tension was reduced. In tadpoles, repeated in vivo observations revealed that PSC processes lead nerve terminal growth. In the absence of PSCs, growth and addition of synapses was dramatically reduced, and existing synapses underwent widespread retraction. Our findings provide in vivo evidence that glial cells maintain presynaptic structure and function at adult synapses and are vital for the growth and stability of developing synapses.