Bcl-2 enhances Ca(2+) signaling to support the intrinsic regenerative capacity of CNS axons

At a certain point in development, axons in the mammalian CNS undergo a profound loss of intrinsic growth capacity, which leads to poor regeneration after injury. Overexpression of Bcl-2 prevents this loss, but the molecular basis of this effect remains unclear. Here, we report that Bcl-2 supports axonal growth by enhancing intracellular Ca(2+) signaling and activating cAMP response element binding protein (CREB) and extracellular-regulated kinase (Erk), which stimulate the regenerative response and neuritogenesis. Expression of Bcl-2 decreases endoplasmic reticulum (ER) Ca(2+) uptake and storage, and thereby leads to a larger intracellular Ca(2+) response induced by Ca(2+) influx or axotomy in Bcl-2-expressing neurons than in control neurons. Bcl-x(L), an antiapoptotic member of the Bcl-2 family that does not affect ER Ca(2+) uptake, supports neuronal survival but cannot activate CREB and Erk or promote axon regeneration. These results suggest a novel role for ER Ca(2+) in the regulation of neuronal response to injury and define a dedicated signaling event through which Bcl-2 supports CNS regeneration.     

Identification and functional expression of a family of nicotinic acetylcholine receptor subunits in the central nervous system of the mollusc Lymnaea stagnalis

We described a family of nicotinic acetylcholine receptor (nAChR) subunits underlying cholinergic transmission in the central nervous system (CNS) of the mollusc Lymnaea stagnalis. By using degenerate PCR cloning, we identified 12 subunits that display a high sequence similarity to nAChR subunits, of which 10 are of the alpha-type, 1 is of the beta-type, and 1 was not classified because of insufficient sequence information. Heterologous expression of identified subunits confirms their capacity to form functional receptors responding to acetylcholine. The alpha-type subunits can be divided into groups that appear to underlie cation-conducting (excitatory) and anion-conducting (inhibitory) channels involved in synaptic cholinergic transmission. The expression of the Lymnaea nAChR subunits, assessed by real time quantitative PCR and in situ hybridization, indicates that it is localized to neurons and widespread in the CNS, with the number and localization of expressing neurons differing considerably between subunit types. At least 10% of the CNS neurons showed detectable nAChR subunit expression. In addition, cholinergic neurons, as indicated by the expression of the vesicular ACh transporter, comprise approximately 10% of the neurons in all ganglia. Together, our data suggested a prominent role for fast cholinergic transmission in the Lymnaea CNS by using a number of neuronal nAChR subtypes comparable with vertebrate species but with a functional complexity that may be much higher.     

Variability of morphologic parameters of individual neurons and neuron clusters in the visceral ganglion of the mollusk Lymnaea stagnalis L

Quantitative investigations on variability of visual morphological parameters in neurons BB1, BB3-5 and clusters BBb have been performed on the ventral side of the visceral ganglion in 20 molluscs Lymnaea stagnalis L. (height of the shell -45.5-1.3 mm, variation coefficient -2.9%) cought in the same pond and on the same day. Statistical analysis of the data obtained is performed by means of the methods applied for analysing small independent samples. A probability character of visual identification and an essential variability are demonstrated for the parameters of the neuron BB1 (diameter, position, colour), of the number of giant and/or large neurons BB3-5 and the cluster BBb neurons. The variation character of the parameters mentioned is asymmetric. It is possible to speak on a greater determinancy of a preferable gradation of the parameters. The interconnection of the variability and the determinancy of the morphological signs with the identificity and functioning of the neuronal elements are discussed. A suggestion is made that identification of the neurons according to both their morphological and functional criteria, as a whole, can have a probability character. The data considered in the work make doubt the invariance of the neurons studied in the mollusc Lymnaea stagnalis L.     

Central representation of internal and external sensory information in the CNS of Helix pomatia L. and Lymnaea stagnalis L

The central representation of intero- and exteroreceptors located in visceral organs and the osphradium were compared in the CNS of Helix pomatia L. (Gastropoda, Stylommatophora) and Lymnaea stagnalis L. (Gastropoda, Basommatophora), two pulmonate snail species inhabiting a terrestrial and anaquatic environment, respectively. Semi-intact preparations were used comprising the CNS connected by the corresponding nerves either to the cardio-renal, respiratory and genital systems or to the osphradium. Spike discharges of central neurons and the nerves were recorded simultaneously. The central representation of intero- and exteroreceptors was found to be distributed throughout the CNS and involved about 300 neurons. The majority of the neurons received sensory information from all the studied visceral organs and the osphradium. Among the neurons responding to intero- and exteroreceptors a multimodal reaction to tactile, chemical and osmotic stimuli prevailed while in the osphradium specific reactions also were demonstrated. Central neurons receiving sensory information from visceral organs and the osphradium form overlapping and reorganizing neural circuits using the same neurons in the regulation of heart activity, respiration or reproduction producing the appropriate behaviour. In the selection of sensory information the firing pattern appears to be the main determining factor as bursting neurons do not receive sensory information. The central representation of intero- and exteroreceptors and its variability can be a model system for cellular studies of motivational state and self-perception.     

Influence of HgCl2 on the osphradial multisensory system of Lymnaea stagnalis L

The osphradial multisensory system of Lymnaea stagnalis L. (Pulmonata, Basommatophora) was used to demonstrate the modulation of chemosensory information both at periphery and central nervous system (CNS) following heavy metal treatments. A semi-intact preparation including osphradium, CNS and the right inner parietal nerve (r.i.p.n.) connecting them was used to record both extracellular activity of nerve and intracellular activity of central neurons receiving information from osphradium. The ion currents of osphradium were recorded using patch-clamp method. The changes in nerve and neuronal activity were expressed by averaging of firing frequency and interspike intervals. The chemosensory function of osphradium was shown by application of L-aspartate, urea, saccharose and stagnant water to its surface. The central neurons reacting to the stimulation ofosphradium were located to visceral, right parietal, pedal and cerebral ganglia of Lymnaea. Both the acute and chronic treatments with HgCl2 damaged the sensory function of osphradium traced on the flow of information from periphery to central neurons. At the same time, mercury chloride modified the synaptic connections of respiratory pattern generators as well as the Ca- and K-dependent ion currents of osphradial neurons. The results proved the multisensory role of osphradium sensing the alterations in the environment and its usefulness in monitoring the effects of pollutants at various level of regulation from chemosensory epithelium to CNS.     

Locomotor rhythms in the pond snail Lymnaea stagnalis: site of origin and neurotransmitter requirements

1. We have found that, in preparations of isolated CNS of the pond snail Lymnaea stagnalis, both serotonin (5HT) and dopamine (DA), as well as their respective precursors, 5HTP and DOPA, are effective in producing fictive intense (muscular) locomotion. 2. Phase-coupled to each of the above pedal rhythms are numerous identifiable pedal neurons including the respiratory interneuron RPeD1, thus suggesting interaction between networks responsible for locomotion and air breathing. 3. The novel DA/DOPA-dependent motor rhythm resembles the 5HT/5HTP-dependent one in terms of activity of identifiable pedal neurons, being however considerably slower than the latter. 4. The results of transection experiments suggest that each of the rhythms is generated by a paired CPG lying entirely within the pedal ganglia.     

Two dopamine receptors: biochemistry, physiology and pharmacology

In 1979, two categories of dopamine (DA) receptors (designated as D-1 and D-2) were identified on the basis of the ability of a limited number of agonists and antagonists to discriminate between these two entities. In the past 5 years agonists and antagonists selective for each category of receptor have been identified. Using these selective drugs it has been possible to attribute the effects of DA upon physiological and biochemical processes to the stimulation of either a D-1 or a D-2 receptor. Thus, DA-induced enhancement of both hormone release from bovine parathyroid gland and firing of neurosecretory cells in the CNS of Lymnaea stagnalis has been attributed to stimulation of a D-1 receptor. Likewise, the DA-induced inhibition of the release of prolactin and alpha-MSH from the pituitary gland, as well as of acetylcholine, DA and beta-endorphin from brain, the DA-induced inhibition of chemo-sensory discharge in rabbit carotid body and the DA-induced hyperpolarization of neurosecretory cells in the CNS of Lymnaea stagnalis have been attributed to stimulation of a D-2 receptor. Independently two categories of DA receptors (designated as DA-1 and DA-2) were identified in the cardiovascular system. Stimulation of a DA-1 receptor increases the vascular cyclic AMP content and causes a relaxation of vascular smooth muscle in renal blood vessels, whereas stimulation of a DA-2 receptor inhibits the release of norepinephrine from certain postganglionic sympathetic neurons. Recent studies with the newly developed drugs discriminating between D-1 and D-2 receptors suggest however that the independently developed schemata for classification of dopamine receptors in either the central nervous and endocrine systems or the cardiovascular system are similar although maybe not completely identical.     

Excitatory actions of propofol and ketamine in the snail Lymnaea stagnalis

This study compares the actions of the intravenous anaesthetics propofol and ketamine on animal behaviour and neuronal activity in the snail Lymnaea stagnalis, particularly in relation to excitatory effects observed clinically. When injected into the whole animal, neither agent induced total anaesthesia. Rather, behavioural activity was enhanced by propofol (10(-5) M) and ketamine (10(-7) M), indicating excitatory effects. When superfused over the isolated central nervous system (CNS), differential effects were produced in two identified neurons, right pedal dorsal 1 (RPeD1) and visceral dorsal 4 (VD4). Resting membrane properties were largely unaffected. However, spike after hyperpolarisation was significantly reduced in RPeD1, but not VD4, with some evidence of increased excitability. In addition, an intrinsic bursting property (post-stimulus burst) in VD4 was altered by propofol (10(-7) M). The results suggest significant excitatory components in the actions of some intravenous anaesthetics, as well as a potential role in modifying excitation and bursting mechanisms in the CNS.