scholarly journals Spontaneous Calcium Oscillations through Differentiation: A Calcium Imaging Analysis of Rat Cochlear Nucleus Neural Stem Cells

Cells ◽  
2021 ◽  
Vol 10 (10) ◽  
pp. 2802
Author(s):  
Johannes Voelker ◽  
Christine Voelker ◽  
Jonas Engert ◽  
Nikolas Goemann ◽  
Rudolf Hagen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Calcium Channels ◽  
Neural Stem Cells ◽  
Calcium Imaging ◽  
Cochlear Nucleus ◽  
Auditory Pathway ◽  
Calcium Oscillations ◽  
Imaging Analysis ◽  
Voltage Dependent ◽  
Voltage Dependent Calcium Channels

Causal therapies for the auditory-pathway and inner-ear diseases are still not yet available for clinical application. Regenerative medicine approaches are discussed and examined as possible therapy options. Neural stem cells could play a role in the regeneration of the auditory pathway. In recent years, neural stem and progenitor cells have been identified in the cochlear nucleus, the second nucleus of the auditory pathway. The current investigation aimed to analyze cell maturation concerning cellular calcium activity. Cochlear nuclei from PND9 CD rats were microscopically dissected and propagated as neurospheres in free-floating cultures in stem-cell medium (Neurobasal, B27, GlutaMAX, EGF, bFGF). After 30 days, the dissociation and plating of these cells took place under withdrawal of the growth factors and the addition of retinoic acid, which induces neural cell differentiation. Calcium imaging analysis with BAPTA-1/Oregon Green was carried out at different times during the differentiation phase. In addition, the influence of different voltage-dependent calcium channels was analyzed through the targeted application of inhibitors of the L-, N-, R- and T-type calcium channels. For this purpose, comparative examinations were performed on CN NSCs, and primary CN neurons. As the cells differentiated, a significant increase in spontaneous neuronal calcium activity was demonstrated. In the differentiation stage, specific frequencies of the spontaneous calcium oscillations were measured in different regions of the individual cells. Initially, the highest frequency of spontaneous calcium oscillations was ascertainable in the maturing somata. Over time, these were overtaken by calcium oscillations in the axons and dendrites. Additionally, in the area of the growth cones, an increasing activity was determined. By inhibiting voltage-dependent calcium channels, their expression and function in the differentiation process were confirmed. A comparable pattern of maturation of these channels was found in CN NSCs and primary CN neurons. The present results show that neural stem cells of the rat cochlear nucleus differentiated not only morphologically but also functionally. Spontaneous calcium activities are of great relevance in terms of neurogenesis and integration into existing neuronal structures. These functional aspects of neurogenesis within the auditory pathway could serve as future targets for the exogenous control of neuronal regeneration.

Effects of N- and L-type calcium channel antagonists on the responses of nociceptive spinal cord neurons to mechanical stimulation of the normal and the inflamed knee joint

1996 ◽  
Vol 76 (6) ◽  
pp. 3740-3749 ◽  
Author(s):  
V. Neugebauer ◽  
H. Vanegas ◽  
J. Nebe ◽  
P. Rumenapp ◽  
H. G. Schaible
Keyword(s):  
Spinal Cord ◽  
Knee Joint ◽  
Calcium Channels ◽  
Mechanical Stimulation ◽  
Topical Administration ◽  
Knee Joints ◽  
Spinal Cord Neurons ◽  
Inflammatory Conditions ◽  
Voltage Dependent ◽  
Voltage Dependent Calcium Channels

1. The present study addresses the involvement of voltage-dependent calcium channels of the N and L type in the spinal processing of innocuous and noxious input from the knee joint, both under normal conditions and under inflammatory conditions in which spinal cord neurons become hyperexcitable. In 30 anesthetized rats, extracellular recordings were performed from single dorsal horn neurons in segments 1–4 of the lumbar spinal cord. All neurons had receptive fields in the ipsilateral knee joint. In 22 rats, an inflammation was induced in the ipsilateral knee joint by kaolin and carrageenan 4–16 h before the recordings. The antagonist at N-type calcium channels, omega-conotoxin GVIA (omega-CTx GVIA), was administered topically in solution to the dorsal surface of the spinal cord at the appropriate spinal segments in 6 rats with normal joints and in 12 rats with inflamed knee joints. The antagonist at L-type channels, nimodipine, was administered topically in 5 rats with normal joints and in 11 rats with inflamed knee joints. In another five rats with inflamed joints, antagonists at L-type calcium channels (diltiazem and nimodipine) and omega-CTx GVIA were administered ionophoretically with multibarrel electrodes close to the neurons recorded. 2. The topical administration of omega-CTx GVIA to the spinal cord reduced the responses to both innocuous and noxious pressure applied to the knee joint in a sample of 11 neurons with input from the normal joint and in a sample of 16 neurons with input from the inflamed joint (hyperexcitable neurons). The responses were decreased to approximately 65% of the predrug values within administration times of 30 min. A similar reduction of the responses to innocuous and noxious pressure was observed when omega-CTx GVIA was administered ionophoretically to nine hyperexcitable neurons. In neurons with input from the normal or the inflamed knee joint, the administration of omega-CTx GVIA led also to a reduction of the responses to innocuous and noxious pressure applied to the noninflamed ankle joint. 3. The topical administration of nimodipine decreased the responses to innocuous and noxious pressure applied to the knee in a sample of 9 neurons with input from the normal joint and in a sample of 16 neurons with input from the inflamed knee joint (hyperexcitable neurons). Within administration times of 30 min, the responses were reduced to approximately 70% of the predrug values. In hyperexcitable neurons, the responses to innocuous and noxious pressure applied to the knee were also decreased during ionophoretic administration of nimodipine (6 neurons) and diltiazem (9 neurons). When the noninflamed ankle was stimulated, the responses to innocuous pressure were reduced neither in neurons with input from the normal knee nor in neurons with input from the inflamed knee, but the responses of hyperexcitable neurons to noxious pressure onto the ankle were reduced. The ionophoretic administration of the agonist at the L-type calcium channel, S(-)-Bay K 8644, enhanced the responses to mechanical stimulation of the knee joint in all 14 hyperexcitable neurons tested. The effect of S(-)-Bay K 8644 was counteracted by both diltiazem (in 6 of 6 neurons) and nimodipine (in 5 of 5 neurons). 4. These data show that antagonists at both the N- and the L-type voltage-dependent calcium channels influence the spinal processing of input from the knee joint. The data suggest, therefore, that voltage-dependent calcium calcium channels of both the N and the L type are important for the sensory functions of the spinal cord. They are involved in the spinal processing of nonnociceptive as well as nociceptive mechanosensory input from the joint, both under normal and inflammatory conditions. The present results show in particular that N- and L-type channels are likely to be involved in the generation of pain evoked by noxious mechanical stimulation in normal tissue as well as in the mechanical hyperalgesia that is usually pres

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