Odor Enrichment Increases Hippocampal Neuron Numbers in Mouse.

The hippocampus and olfactory bulb incorporate new neurons migrating from neurogenic regions in the brain. Hippocampal atrophy is evident in numerous neurodegenerative disorders, and altered hippocampal neurogenesis is an early pathological event in Alzheimer's disease. We hypothesized that hippocampal neurogenesis is affected by olfactory stimuli through the neural pathway of olfaction-related memory. In this study, we exposed mice to novel pleasant odors for three weeks and then assessed the number of neurons, non-neuronal cells (mainly glia) and proliferating cells in the hippocampus and olfactory bulb, using the isotropic fractionator method. We found that the odor enrichment significantly increased the neuronal cell numbers in the hippocampus, and promoted cell proliferation and neurogenesis in the olfactory bulb. In contrast, the glial cell numbers remained unchanged in both of the regions. Our results suggest that exposure to novel odor stimuli promotes hippocampal neurogenesis and support the idea that enriched environments may delay the onset or slow down the progression of neurodegenerative disorders.

Individual variability of venom from the European adder (Vipera berus berus) from one locality in Eastern Hungary.

We have revealed intra-population variability among venom samples from several individual European adders (Vipera berus berus) within a defined population in Eastern Hungary. Individual differences in venom pattern were noticed, both gender-specific and age-related, by one-dimensional electrophoresis. Gelatin zymography demonstrated that these individual venoms have different degradation profiles indicating varying protease activity in the specimens from adders of different ages and genders. Some specimens shared a conserved region of substrate degradation, while others had lower or extremely low protease activity. Phospholipase A2 activity of venoms was similar but not identical. Interspecimen diversity of the venom phospholipase A2-spectra (based on the components' molecular masses) was detected by MALDI-TOF MS. The lethal toxicity of venoms (LD50) also showed differences among individual snakes. Extracted venom samples had varying neuromuscular paralysing effect on chick biventer cervicis nerve-muscle preparations. The paralysing effect of venom was lost when calcium in the physiological salt solution was replaced by strontium; indicating that the block of twitch responses to nerve stimulation is associated with the activity of a phospholipase-dependent neurotoxin. In contrast to the studied V. b. berus venoms from different geographical regions so far, this is the first V. b. berus population discovered to have predominantly neurotoxic neuromuscular activity. The relevance of varying venom yields is also discussed. This study demonstrates that individual venom variation among V. b. berus living in particular area of Eastern Hungary might contribute to a wider range of clinical manifestations of V. b. berus envenoming than elsewhere in Europe.

Adult Neurogenesis and Gliogenesis: Possible Mechanisms for Neurorestoration.

The subgranular zone (SGZ) and subventricular zone (SVZ) are developmental remnants of the germinal regions of the brain, hence they retain the ability to generate neuronal progenitor cells in adult life. Neurogenesis in adult brain has an adaptive function because newly produced neurons can integrate into and modify existing neuronal circuits. In contrast to the SGZ and SVZ, other brain regions have a lower capacity to produce new neurons, and this usually occurs via parenchymal and periventricular cell genesis. Compared to neurogenesis, gliogenesis occurs more prevalently in the adult mammalian brain. Under certain circumstances, interaction occurs between neurogenesis and gliogenesis, facilitating glial cells to transform into neuronal lineage. Therefore, modulating the balance between neurogenesis and gliogenesis may present a new perspective for neurorestoration, especially in diseases associated with altered neurogenesis and/or gliogenesis, cell loss, or disturbed homeostasis of cellular constitution. The present review discusses important neuroanatomical features of adult neurogenesis and gliogenesis, aiming to explore how these processes could be modulated toward functional repair of the adult brain.

Early in vivo Effects of the Human Mutant Amyloid-ß Protein Precursor (hAßPPSwInd) on the Mouse Olfactory Bulb.

The amyloid-ß protein precursor (AßPP) has long been linked to Alzheimer's disease (AD). Using J20 mice, which express human AßPP with Swedish and Indiana mutations, we studied early pathological changes in the olfactory bulb. The presence of AßPP/amyloid-ß (Aß) was examined in mice aged 3 months (before the onset of hippocampal Aß deposition) and over 5 months (when hippocampal Aß deposits are present). The number of neurons, non-neurons, and proliferating cells was assessed using the isotropic fractionator method. Our results demonstrate that although AßPP is overexpressed in some of the mitral cells, widespread Aß deposition and microglia aggregates are not prevalent in the olfactory bulb. The olfactory bulbs of the younger J20 group harbored significantly fewer neurons than those of the age-matched wild-type mice (5.57±0.13 million versus 6.59±0.36 million neurons; p = 0.011). In contrast, the number of proliferating cells was higher in the young J20 than in the wild-type group (i.e., 6617±425 versus 4455±623 cells; p = 0.011). A significant increase in neurogenic activity was also observed in the younger J20 olfactory bulb. In conclusion, our results indicate that (1) neurons participating in the mouse olfactory function overexpress AßPP; (2) the cellular composition of the young J20 olfactory bulb is different from that of wild-type littermates; (3) these differences may reflect altered neurogenic activity and/or delayed development of the J20 olfactory system; and (4) AßPP/Aß-associated pathological changes that take place in the J20 hippocampus and olfactory bulb are not identical.

The spinal cord of the common marmoset (Callithrix jacchus).

The marmoset spinal cord possesses all the characteristic features of a typical mammalian spinal cord, but with some interesting variation in the levels of origin of the limb nerves. In our study Nissl and ChAT sections of the each segment of the spinal cord in two marmosets (Ma5 and Ma8), we found that the spinal cord can be functionally and anatomically divided into six regions: the prebrachial region (C1 to C3); the brachial region (C4 to C8) - segments supplying the upper limb; the post-brachial region (T1 to L1) - containing the sympathetic outflow, and supplying the hypaxial muscles of the body wall; the crural region (L2 to L5) - segments supplying the lower limb; the postcrural region (L6) - containing the parasympathetic outflow; and the caudal region (L7 to Co4) - supplying the tail. In the rat, mouse, and rhesus monkey, the prebrachial region consists of segments C1 to C4 (with the phrenic nucleus located at the C4 segment), and the brachial region extends from C5 to T1 inclusive. The prefixing of the upper limb outflow in these two marmosets mirrors the finding in the literature that a large C4 contribution to the brachial plexus is common in humans.

Silencing the KCNK9 potassium channel (TASK-3) gene disturbs mitochondrial function, causes mitochondrial depolarization, and induces apoptosis of human melanoma cells.

TASK-3 (KCNK9 or K2P9.1) channels are thought to promote proliferation and/or survival of malignantly transformed cells, most likely by increasing their hypoxia tolerance. Based on our previous results that suggested mitochondrial expression of TASK-3 channels, we hypothesized that TASK-3 channels have roles in maintaining mitochondrial activity. In the present work we studied the effect of reduced TASK-3 expression on the mitochondrial function and survival of WM35 and A2058 melanoma cells. TASK-3 knockdown cells had depolarized mitochondrial membrane potential and contained a reduced amount of mitochondrial DNA. Compared to their scrambled shRNA-transfected counterparts, they demonstrated diminished responsiveness to the application of the mitochondrial uncoupler [(3-chlorophenyl)hydrazono]malononitrile (CCCP). These observations indicate impaired mitochondrial function. Further, TASK-3 knockdown cells presented reduced viability, decreased total DNA content, altered cell morphology, and reduced surface area. In contrast to non- and scrambled shRNA-transfected melanoma cell lines, which did not present noteworthy apoptotic activity, almost 50 % of the TASK-3 knockdown cells exhibited strong Annexin-V-specific immunofluorescence signal. Sequestration of cytochrome c from the mitochondria to the cytosol, increased caspase 3 activity, and translocation of the apoptosis-inducing factor from mitochondria to cell nuclei were also demonstrated in TASK-3 knockdown cells. Interference with TASK-3 channel expression, therefore, induces caspase-dependent and -independent apoptosis of melanoma cells, most likely via causing mitochondrial depolarization. Consequently, TASK-3 channels may be legitimate targets of future melanoma therapies.

Age-dependent alterations of the hippocampal cell composition and proliferative potential in the hAßPPSwInd-J20 mouse.

The J20 mouse expresses human mutant amyloid-ß protein precursor (hAßPPSwInd) and is an established transgenic model of Alzheimer's disease (AD). From the age of 5 months, amyloid-ß (Aß) deposits appear in the hippocampus with concomitant increase of AD-associated features. Although changes occurring after the appearance of Aß deposits have been extensively studied, very little is known about alterations that occur prior to 5 months. The present study aimed to identify changes in the cellular composition and proliferative potential of the J20 hippocampus using 1-18-month-old mice. Neuronal, non-neuronal, Ki-67+, and TUNEL+ cell numbers were counted with the isotropic fractionator method. Age-dependent changes of the expression of microglia-, astrocyte-, and neurogenesis-specific markers were sought in the entire hippocampus. Several transgene-associated changes were revealed before the appearance of Aß deposits. The number of proliferating cells decreased whereas the number of microglia clusters increased as early as 4 weeks of age. The neurogenesis was also impaired in the dentate gyrus of 7-11-week-old J20 mice. A statistically significant negative correlation was found between the number of proliferating cells and age in both populations, but the time course of the age-dependence was steeper in wild-type than in J20 mice. Negative age-dependence was noted when the number of cells committed to apoptosis was examined. Our results indicate that overexpression of mutant hAßPP initiates a cascade of pathologic events well before the appearance of visible Aß plaques. Accordingly, early signs of AD include reduced cell proliferation, impaired neurogenesis, and increased activity of microglia in the hippocampus.

Musculotopic organization of the motor neurons supplying the mouse hindlimb muscles: a quantitative study using Fluoro-Gold retrograde tracing.

We have mapped the motor neurons (MNs) supplying the major hindlimb muscles of transgenic (C57/BL6J-ChAT-EGFP) and wild-type (C57/BL6J) mice. The fluorescent retrograde tracer Fluoro-Gold was injected into 19 hindlimb muscles. Consecutive transverse spinal cord sections were harvested, the MNs counted, and the MN columns reconstructed in 3D. Three longitudinal MN columns were identified. The dorsolateral column extends from L4 to L6 and consists of MNs innervating the crural muscles and the foot. The ventrolateral column extends from L1 to L6 and accommodates MNs supplying the iliopsoas, gluteal, and quadriceps femoris muscles. The middle part of the ventral horn hosts the central MN column, which extends between L2 and L6 and consists of MNs for the thigh adductor, hamstring, and quadratus femoris muscles. Within these longitudinal columns, the arrangement of the different MN groups reflects their somatotopic organization. MNs innervating muscles developing from the dorsal (e.g., quadriceps) and ventral muscle mass (e.g., hamstring) are situated in the lateral and medial part of the ventral gray, respectively. MN pools belonging to proximal muscles (e.g., quadratus femoris and iliopsoas) are situated ventral to those supplying more distal ones (e.g., plantar muscles). Finally, MNs innervating flexors (e.g., posterior crural muscles) are more medial than those belonging to extensors of the same joint (e.g., anterior crural muscles). These data extend and modify the MN maps in the recently published atlas of the mouse spinal cord and may help when assessing neuronal loss associated with MN diseases.

The hyperpolarization-activated non-specific cation current (In ) adjusts the membrane properties, excitability, and activity pattern of the giant cells in the rat dorsal cochlear nucleus.

Giant cells of the cochlear nucleus are thought to integrate multimodal sensory inputs and participate in monaural sound source localization. Our aim was to explore the significance of a hyperpolarization-activated current in determining the activity of giant neurones in slices prepared from 10 to 14-day-old rats. When subjected to hyperpolarizing stimuli, giant cells produced a 4-(N-ethyl-N-phenylamino)-1,2-dimethyl-6-(methylamino) pyridinium chloride (ZD7288)-sensitive inward current with a reversal potential and half-activation voltage of -36 and -88 mV, respectively. Consequently, the current was identified as the hyperpolarization-activated non-specific cationic current (Ih ). At the resting membrane potential, 3.5% of the maximum Ih conductance was available. Immunohistochemistry experiments suggested that hyperpolarization-activated, cyclic nucleotide-gated, cation non-selective (HCN)1, HCN2, and HCN4 subunits contribute to the assembly of the functional channels. Inhibition of Ih hyperpolarized the membrane by 6 mV and impeded spontaneous firing. The frequencies of spontaneous inhibitory and excitatory postsynaptic currents reaching the giant cell bodies were reduced but no significant change was observed when evoked postsynaptic currents were recorded. Giant cells are affected by biphasic postsynaptic currents consisting of an excitatory and a subsequent inhibitory component. Inhibition of Ih reduced the frequency of these biphasic events by 65% and increased the decay time constants of the inhibitory component. We conclude that Ih adjusts the resting membrane potential, contributes to spontaneous action potential firing, and may participate in the dendritic integration of the synaptic inputs of the giant neurones. Because its amplitude was higher in young than in adult rats, Ih of the giant cells may be especially important during the postnatal maturation of the auditory system.

Musculotopic organization of the motor neurons supplying forelimb and shoulder girdle muscles in the mouse.

We identified the motor neurons (MNs) supplying the shoulder girdle and forelimb muscles in the C57BL/6J mouse spinal cord using Fluoro-Gold retrograde tracer injections. In spinal cord transverse sections from C2 to T2, we observed two MN columns (medial and lateral) both with ventral and dorsal subdivisions. The dorsolateral column consisted of the biceps brachii, forearm extensors, forearm flexors, and hand MNs, and the ventrolateral column consisted of the latissimus dorsi, trapezius, teres major, deltoid, and triceps MNs. The supraspinatus muscle MNs were located in the dorsomedial column, and pectoralis major and serratus anterior MNs were located in the ventromedial columns. MNs of the dorsolateral column innervated the biceps brachii in mid-C4 to mid-C7, forearm extensors in caudal C4 to mid-T1, forearm flexors in rostral C5 to mid-T1, and hand muscles in mid-C8 to mid-T2 segments. The MNs innervating the trapezius were located in mid-C2 to mid-C4, triceps brachii in mid-C6 to rostral T1, deltoid in rostral C4 to mid-C6, teres major in rostral C5 to mid-C8, and latissimus dorsi in mid-C5 to caudal C8. In addition, MNs innervating the supraspinatus were located from rostral C4 to caudal C8, pectoralis major in mid-C6 to mid-T2, and serratus anterior in rostral C5 to caudal C7/rostral C8 segments. While the musculotopic pattern of MN groups was very similar to that documented for other species, we found differences in the position and cranio-caudal extent of some MN pools compared with previous reports. The identification of mouse forelimb MNs can serve as an anatomical reference for studying degenerative MN diseases, spinal cord injury, and developmental gene expression.

Three-dimensional reconstruction and quantitative morphometric analysis of pyramidal and giant neurons of the rat dorsal cochlear nucleus.

Correct interpretation of functional data obtained from various cell types of the cochlear nucleus (CN), a structure involved in auditory information processing, necessitates reliable cell identification. Our aim was to perform a quantitative morphological characterization of giant and pyramidal cells of the rat CN and identify parameters that are suitable for their adequate classification. Neurons were labeled with biocytin, visualized with a fluorescent marker, and three-dimensionally reconstructed from confocal images. The size and shape of the soma and dendritic tree of each neuron were characterized by 17 morphometric parameters. The variables were subjected to multivariate statistical analysis to determine their importance while discriminating between giant and pyramidal cells. Our results provide a new battery of morphometric data, which could not be obtained earlier, improve the chances of correct cell identification, make modeling experiments easier and more reliable, and help us to understand both the functions of individual CN neurons and the network properties of this nucleus. In addition, we demonstrate that even partial labeling and/or incomplete reconstruction of neurons may be enough for their correct identification if selected parameters describing the cell bodies and the proximal portions of the dendritic trees are utilized. We propose that our findings have specific relevance to studies which attempt cell identification after functional experiments resulting in incomplete labeling of the investigated neurons.

Cellular composition characterizing postnatal development and maturation of the mouse brain and spinal cord.

The process of development, maturation, and regression in the central nervous system (CNS) are genetically programmed and influenced by environment. Hitherto, most research efforts have focused on either the early development of the CNS or the late changes associated with aging, whereas an important period corresponding to adolescence has been overlooked. In this study, we searched for age-dependent changes in the number of cells that compose the CNS (divided into isocortex, hippocampus, olfactory bulb, cerebellum, 'rest of the brain', and spinal cord) and the pituitary gland in 4-40-week-old C57BL6 mice, using the isotropic fractionator method in combination with neuronal nuclear protein as a marker for neuronal cells. We found that all CNS structures, except for the isocortex, increased in mass in the period of 4-15 weeks. Over the same period, the absolute number of neurons significantly increased in the olfactory bulb and cerebellum while non-neuronal cell numbers increased in the 'rest of the brain' and isocortex. Along with the gain in body length and weight, the pituitary gland also increased in mass and cell number, the latter correlating well with changes of the brain and spinal cord mass. The majority of the age-dependent alterations (e.g., somatic parameters, relative brain mass, number of pituitary cells, and cellular composition of the cerebellum, isocortex, rest of the brain, and spinal cord) occur rapidly between the 4th and 11th postnatal weeks. This period includes murine adolescence, underscoring the significance of this stage in the postnatal development of the mouse CNS.

Protein phosphatase-1M and Rho-kinase affect exocytosis from cortical synaptosomes and influence neurotransmission at a glutamatergic giant synapse of the rat auditory system.

Protein phosphatase-1M (PP1M, myosin phosphatase) consists of a PP1 catalytic subunit (PP1c) and the myosin phosphatase target subunit-1 (MYPT1). RhoA-activated kinase (ROK) regulates PP1M via inhibitory phosphorylation of MYPT1. Using multidisciplinary approaches, we have studied the roles of PP1M and ROK in neurotransmission. Electron microscopy demonstrated the presence of MYPT1 and ROK in both pre- and post-synaptic terminals. Tautomycetin (TMC), a PP1-specific inhibitor, decreased the depolarization-induced exocytosis from cortical synaptosomes. trans-4-[(1R)-1-aminoethyl]-N-4-pyridinylcyclohexanecarboxamide dihydrochloride, a ROK-specific inhibitor, had the opposite effect. Mass spectrometry analysis identified several MYPT1-bound synaptosomal proteins, of which interactions of synapsin-I, syntaxin-1, calcineurin-A subunit, and Ca(2+) /calmodulin-dependent kinase II with MYPT1 were confirmed. In intact synaptosomes, TMC increased, whereas Y27632 decreased the phosphorylation levels of MYPT1(Thr696) , myosin-II light chain(Ser19) , synapsin-I(Ser9) , and syntaxin-1(Ser14) , indicating that PP1M and ROK influence their phosphorylation status. Confocal microscopy indicated that MYPT1 and ROK are present in the rat ventral cochlear nucleus both pre- and post-synaptically. Analysis of the neurotransmission in an auditory glutamatergic giant synapse demonstrated that PP1M and ROK affect neurotransmission via both pre- and post-synaptic mechanisms. Our data suggest that both PP1M and ROK influence synaptic transmission, but further studies are needed to give a full account of their mechanism of action.

Activation of muscarinic receptors increases the activity of the granule neurones of the rat dorsal cochlear nucleus--a calcium imaging study.

Acetylcholine modulates the function of the cochlear nucleus via several pathways. In this study, the effects of cholinergic stimulation were studied on the cytoplasmic Ca(2+) concentration of granule neurones of the rat dorsal cochlear nucleus (DCN). Ca(2+) transients were recorded in Oregon-Green-BAPTA 1-loaded brain slices using a calcium imaging technique. For the detection, identification and characterisation of the Ca(2+) transients, a wavelet analysis-based method was developed. Granule cells were identified on the basis of their size and localisation. The action potential-coupled character of the Ca(2+) transients of the granule cells was established by recording fluorescence changes and electrical activity simultaneously. Application of the cholinergic agonist carbamyl-choline (CCh) significantly increased the frequency of the Ca(2+) transients (from 0.37 to 6.31 min(-1), corresponding to a 17.1-fold increase; n = 89). This effect was antagonised by atropine, whereas CCh could still evoke an 8.3-fold increase of the frequency of the Ca(2+) transients when hexamethonium was present. Using immunolabelling, the expression of both type 1 and type 3 muscarinic receptors (M1 and M3 receptors, respectively) was demonstrated in the granule cells. Application of 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide (an M3-specific antagonist) prevented the onset of the CCh effect, whereas an M1-specific antagonist (pirenzepine) was less effective. We conclude that cholinergic stimulation increases the activity of granule cells, mainly by acting on their M3 receptors. The modulation of the firing activity of the granule cells, in turn, may modify the firing of projection neurones and may adjust signal processing in the entire DCN.

A cytoarchitectonic and chemoarchitectonic analysis of the dopamine cell groups in the substantia nigra, ventral tegmental area, and retrorubral field in the mouse.

The three main dopamine cell groups of the brain are located in the substantia nigra (A9), ventral tegmental area (A10), and retrorubral field (A8). Several subdivisions of these cell groups have been identified in rats and humans but have not been well described in mice, despite the increasing use of mice in neurodegenerative models designed to selectively damage A9 dopamine neurons. The aim of this study was to determine whether typical subdivisions of these dopamine cell groups are present in mice. The dopamine neuron groups were analysed in 15 adult C57BL/6J mice by anatomically localising tyrosine hydroxylase (TH), dopamine transporter protein (DAT), calbindin, and the G-protein-activated inward rectifier potassium channel 2 (GIRK2) proteins. Measurements of the labeling intensity, neuronal morphology, and the proportion of neurons double-labeled with TH, DAT, calbindin, or GIRK2 were used to differentiate subregions. Coronal maps were prepared and reconstructed in 3D. The A8 cell group had the largest dopamine neurons. Five subregions of A9 were identified: the reticular part with few dopamine neurons, the larger dorsal and smaller ventral dopamine tiers, and the medial and lateral parts of A9. The latter has groups containing some calbindin-immunoreactive dopamine neurons. The greatest diversity of dopamine cell types was identified in the seven subregions of A10. The main dopamine cell groups in the mouse brain are similar in terms of diversity to those observed in rats and humans. These findings are relevant to models using mice to analyse the selective vulnerability of different types of dopamine neurons.

Inhibition of TASK-3 (KCNK9) channel biosynthesis changes cell morphology and decreases both DNA content and mitochondrial function of melanoma cells maintained in cell culture.

TASK-3 channel overexpression was shown to facilitate the survival of malignantly transformed cells, possibly by providing greater hypoxia tolerance through a still unknown mechanism. Although it has been suggested previously that TASK-3 channels are expressed in the mitochondrial membranes, their role here remains elusive. In this study, a transient transfection of TASK-3 knockdown melanoma cell cultures was produced to show the significance of TASK-3 expression. Reduction of the TASK-3 protein biosynthesis induced characteristic changes in cell morphology, reduced the amount of DNA and decreased metabolic activity and mitochondrial function of melanoma cells when compared with control. These findings indicate that TASK-3 channel expression and function is indispensable for the proliferation and/or survival of the melanoma cells, as they seem to contribute to their mitochondrial functions. The significance is that, in this study, we have shown that TASK-3 channels are expressed in the mitochondria of melanoma malignum cells, and they are essential for maintaining cellular integrity and viability. The TASK-3 knockdown melanoma cell line had altered morphology, reduced DNA content, decreased metabolic activity and impaired mitochondrial function. These data indicate that TASK-3 channels are functionally present in the mitochondria of the melanoma cells, and their function is essential for the survival of these cells, thus TASK-3 channels may be the possible targets of future anticancer therapy.

Cytoplasmic Ca2+ concentration changes evoked by muscarinic cholinergic stimulation in primary and metastatic melanoma cell lines.

Experiments were performed to explore differences between cultured primary and metastatic melanoma cell lines in their muscarinic acetylcholine receptor-mediated intracellular Ca signalization. The expression of type 1 and type 3 muscarinic receptors was detected and compared at the protein level using both immunocytochemistry and semiquantitative western blotting. The functionality of muscarinic receptors was tested by applying carbamylcholine (CCh; 1 mmol/l) and by recording the associated increases in cytoplasmic Ca using Ca imaging with the application of the Ca indicator dye, fluo-4. These data indicate that the expression levels of the receptor proteins were not significantly different in the metastatic (HT199, HT168-M1) and the primary (WM35) cell lines. Although Ca transients were evoked in all the three cell lines by CCh, the proportion of the CCh-positive cells was smaller amongst the WM35 cells. The Ca transients could be effectively blocked by atropine (0.1 mmol/l). The time courses of the Ca transients were highly variable, and in some instances they showed a late (plateau-like) component whose presence crucially depended on the influx of extracellular Ca. When the extracellular Ca concentration was reduced, the duration of the CCh-evoked transients was considerably decreased; a phenomenon that was more pronounced in the metastatic cell lines. Although there are no fundamental differences in the muscarinic receptor-mediated Ca signalization of the primary and metastatic cell lines, the quantitative differences showed in this study may partially explain the increased malignancy and migratory potential of the metastatic cells.

Purkinje-like cells of the rat cochlear nucleus: a combined functional and morphological study.

Purkinje-like cells (PLCs) of the cochlear nucleus (CN) are strongly calbindin positive neurones with unknown function. In the present work functional and morphological methods have been employed to provide data about PLCs in general, and about their possible involvement in the synaptic organisation of the CN in particular. PLCs had slightly elongated soma, from which a complex dendritic arborisation extended with highly variable dimensions. On the basis of their morphology, three classes of PLCs were identified. Positively identified PLCs fired a train of action potentials on sustained depolarization. When hyperpolarizing stimuli were applied, the presence of a slowly activating, ZD7288-sensitive inward current was noted that corresponded to the h-current. PLCs received both excitatory and inhibitory synaptic inputs. Functional experiments revealed that 76% and 14% of the spontaneous inhibitory postsynaptic currents recorded from the cell bodies of the PLCs were mediated via glycinergic and GABAergic synapses, respectively. PLCs presented strong cerebellin1-like immunoreactivity, but its distribution differed from that seen in cerebellar Purkinje cells. Our results indicate that PLCs are parts of the synaptic circuitry of the CN, thus they may be actively involved in the processing and analysis of auditory information.

Cytoplasmic Ca(2+) concentration changes evoked by cholinergic stimulation in primary astrocyte cultures prepared from the rat cochlear nucleus.

The involvement of astrocytes in the cholinergic modulation of the cochlear nucleus has been studied using primary astrocyte cultures prepared from this nucleus. The cells were loaded with the membrane permeable form of the fluorescent Ca(2+) indicator Fluo-4, and carbachol-induced Ca(2+) concentration increases were monitored using an imaging system. In the presence of cholinergic stimulation 36.3% of the cells produced Ca(2+) transients. The time course of the transients was variable; 45.0% of the responding cells showed only a rapid Ca(2+) concentration increase, while in 50.5% of the astrocytes the fast component was followed by a slow plateau phase. Using muscarine as well as general and more specific cholinergic antagonists (atropine, pirenzepine, 4-DAMP and hexamethonium), the role of the M3 and (to a smaller extent) M1 muscarinic acetylcholine receptors could be demonstrated in the genesis of the carbachol-induced Ca(2+) transients. The presence of these two subtypes of muscarinic receptors has been confirmed at both mRNA (Q-PCR) and protein (immunocytochemistry) levels. Our data demonstrate the responsiveness of the cochlear astrocytes towards cholinergic stimulation, suggesting that they may have roles in mediating the effects of cholinergic modulation in the rat cochlear nucleus.

Spiral ganglion neurones: an overview of morphology, firing behaviour, ionic channels and function.

The spiral ganglion cells provide the afferent innervation of the hair cells of the organ of Corti. Ninety-five percent of these cells (termed type I spiral ganglion neurones) are in synaptic contact with the inner hair cells, whereas about 5% of them are type II cells, which are responsible for the sensory innervation of the outer hair cells. To understand the function of the spiral ganglion neurones, it is important to explore their membrane properties, understand their activity patterns and describe the variety of ionic channels determining their behaviour. In this review, a brief description is given of the various experimental methods that allow the investigation of the spiral ganglion cells, followed by the discussion of their action potential firing patterns and ionic conductances. The presence, distribution and significance of the K(+) currents of the spiral ganglion cells are specifically addressed, along with the introduction of the putative subunit compositions of the relevant voltage-gated K(+) channels.