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PubMed1998

Effects of age at onset of production, light regime and dietary calcium on performance, eggshell traits, duodenal calbindin and cholecalciferol metabolism.

1. Rate of production and shell thickness (ST) decreased, while body weight (BW), egg weight (EW) and percentage breakage increased progressively with age. Shell weight (SW) increased until 8 to 13 months of age and then decreased. 2. Early onset of production resulted in lower BW and EW at the onset of production, and lower pooled averages of BW, EW, SW and ST, as compared with late or medial onset of production. In 4 out of 5 trials, early onset did not result in the production of more eggs during the laying period. 3. Early onset of production is associated with physiological Ca deficiency as indicated by increases in kidney-1-hydroxylase and duodenal calbindin in early layers as compared with late layers. Early layers exhibited a more severe reduction in shell quality as the result of Ca deficiency as compared with late layers. 4. Feeding pullets with a prelaying diet containing 3.9% Ca did not affect unequivocally the performance or shell quality during the whole productive period, whether the birds started to lay early or late. The dietary treatment did not cause renal damage, as indicated by morphological examination and by plasma calcium and uric acid concentration.

Age Factors↗

PubMed1994

Induction of a calbindin-D9K-like protein in avian muscle cells by 1,25-dihydroxy-vitamin D3.

1,25-dihydroxy-vitamin D3 induces the synthesis of a 9 kDa calcium binding protein in chick embryo myoblasts. This work revealed comigration of the myoblast protein with rat calbindin-D9K in SDS-polyacrylamide gels. Western-blot analysis with a specific calbindin-D9K antibody showed the presence of an immunoreactive protein of 9 kDa in chick tissues. In addition, higher levels of myoblast mRNA hybridizing to a specific cDNA probe for rat calbindin-D9K were detected in response to 1,25-dihydroxy-vitamin D3. Northern hybridization analysis showed that the increase was related to a single 550 nucleotide mRNA species. These results provide the first evidence on the presence of calbindin-D9K or a closely related protein in avian tissues and its inducible expression in muscle by 1,25-dihydroxy-vitamin D3.

Animals↗

PubMed2006

Axotomy induces contrasting changes in calcium and calcium-binding proteins in oculomotor and hypoglossal nuclei of Balb/c mice.

Motor neurons with different susceptibility to degeneration have been identified in amyotrophic lateral sclerosis (ALS). Increase of intracellular calcium has been proposed as a mediator, amplifying the damage through a positive feedback of the known pathological processes. Accordingly, the potential of motor neurons to limit calcium increases during injury might be proportional to their viability. A basic mechanism of reducing calcium amplitudes depends on the calcium-buffering capacity, determined by the calcium-binding protein content. In this study, oculomotor and hypoglossal neurons, prototypes of resistant and vulnerable motor neurons in ALS were examined in axotomy experiments. Total calcium-, parvalbumin-, and calbindin-D28k levels of motor neurons of adult mice were characterized by electron microscopic histochemistry and light microscopic immunostaining. In hypoglossal neurons, compared with oculomotor neurons, larger and more enduring increases of calcium were detected. The perikarya of hypoglossal neurons remained immunonegative for both parvalbumin and calbindin-D28k. Qualitatively, no major cell loss was noted after axotomy, but a decreased neuronal marker staining at days 1-14 suggested a reversible injury of hypoglossal neurons. Oculomotor neurons were not stained for calbindin-D28k but stained for parvalbumin in control conditions, staining which increased at postoperative days 7-14 before returning to baseline. Neuronal marker staining did not change in these cells during the observed period. The higher level of parvalbumin in resistant motor neurons and their ability to up-regulate parvalbumin after injury, paralleled by a smaller increase of intracellular calcium suggest that parvalbumin may have a protective effect in these cells.

Animals↗

PubMed2006

Critical role of the epithelial Ca2+ channel TRPV5 in active Ca2+ reabsorption as revealed by TRPV5/calbindin-D28K knockout mice.

The epithelial Ca(2+) channel TRPV5 facilitates apical Ca(2+) entry during active Ca(2+) reabsorption in the distal convoluted tubule. In this process, cytosolic Ca(2+) remains at low nontoxic concentrations because the Ca(2+) influx is buffered rapidly by calbindin-D(28K). Subsequently, Ca(2+) that is bound to calbindin-D(28K) is shuttled toward the basolateral Ca(2+) extrusion systems. For addressing the in vivo role of TRPV5 and calbindin-D(28K) in the maintenance of the Ca(2+) balance, single- and double-knockout mice of TRPV5 and calbindin-D(28K) (TRPV5(-/-), calbindin-D(28K)(-/-), and TRPV5(-/-)/calbindin-D(28K)(-/-)) were characterized. These mice strains were fed two Ca(2+) diets (0.02 and 2% wt/wt) to investigate the influence of dietary Ca(2+) content on the Ca(2+) balance. Urine analysis indicated that TRPV5(-/-)/calbindin-D(28K)(-/-) mice exhibit on both diets hypercalciuria compared with wild-type mice. Ca(2+) excretion in TRPV5(-/-)/calbindin-D(28K)(-/-) mice was not significantly different from TRPV5(-/-) mice, whereas calbindin-D(28K)(-/-) mice did not show hypercalciuria. The similarity between TRPV5(-/-)/calbindin-D(28K)(-/-) and TRPV5(-/-) mice was supported further by an equivalent increase in renal calbindin-D(9K) expression and in intestinal Ca(2+) hyperabsorption as a result of upregulation of calbindin-D(9K) and TRPV6 expression in the duodenum. Elevated serum parathyroid hormone and 1,25-dihydroxyvitamin D(3) levels accompanied the enhanced expression of the Ca(2+) transporters. Intestinal Ca(2+) absorption and expression of calbindin-D(9K) and TRPV6, as well as serum parameters of the calbindin-D(28K)(-/-) mice, did not differ from those of wild-type mice. These results underline the gatekeeper function of TRPV5 being the rate-limiting step in active Ca(2+) reabsorption, unlike calbindin-D(28K), which possibly is compensated by calbindin-D(9K).

Absorption↗

PubMed1995

Effects of presence of an egg and calcium deposition in the shell gland on levels of messenger ribonucleic acid of CaBP-D28K and of vitamin D3 receptor in the shell gland of the laying hen.

Levels of calbindin-D28K (CaBP-D28K) mRNA and vitamin D3, 1 alpha, 25(OH)2D3, receptor (VDR) mRNA in the intestine and shell gland were measured in the laying hen during the ovulatory cycle by Northern and slot blot analyses. In addition, effects of presence of an egg and calcium deposition in the shell gland on the levels of both mRNAs were studied either by delay in oviposition (retaining the egg in the shell gland beyond the expected time of oviposition) or premature oviposition (emptying the shell gland and arrest of calcium deposition). During the ovulatory cycle mRNA levels of CaBP-D28K and of VDR remained relatively constant in the intestine. In contrast, both mRNA levels of the shell gland were low when there was an egg in the infundibulum, magnum, and isthmus, but significantly increased when there was an egg in the shell gland during shell formation. The im injection of indomethacin 3 hr before expected ovulation delayed oviposition but increased neither shell thickness nor mRNA levels of CaBP-D28K and of VDR. On the other hand, premature oviposition reduced mRNA levels of both CaBP-D28K and VDR in the shell gland. These results suggest that the presence of an egg and calcium deposition in the shell gland may be a stimulatory factor for synthesis and accumulation of CaBP-D28K mRNA and VDR mRNA in association with calcification.

Animals↗

PubMed1991

Induction of calbindin-D 9k mRNA but not calcium transport in rat intestine by 1,25-dihydroxyvitamin D3 24-homologs.

The function and precise mechanism of regulation of calbindin-D 9k in intestine is largely unknown. It is suggested that this calcium binding protein is involved in active intestinal calcium transport and that its expression is mainly mediated by 1,25-dihydroxyvitamin D3. We examined the effect of two side chain modified analogs of 1,25-dihydroxyvitamin D3 as compared to 1,25-dihydroxyvitamin D3 itself on the regulation of the calbindin-D 9k at the mRNA level and on intestinal calcium transport in the rat. delta 22-24,24-dihomo-1,25-dihydroxyvitamin D3 at a single dose of 500, 1,000, and 2,000 pmol caused greater than 7.0-fold increase in calbindin-D 9k mRNA without stimulating intestinal calcium transport. A 10,000-pmol dose of delta 22-24,24,24-trihomo-1,25-dihydroxyvitamin D3 caused a 7.6-fold increase in calbindin-D 9k mRNA without significantly increasing intestinal absorption of calcium. In contrast, 1,25-dihydroxyvitamin D3 caused a parallel increase in calbindin-D 9k mRNA and intestinal absorption of calcium. Thus, calbindin 9k is not by itself responsible for 1,25-dihydroxyvitamin D3-mediated increase in intestinal absorption of calcium.

Animals↗

PubMed1999

Expression of calbindin-D28k in C6 glial cells stabilizes intracellular calcium levels and protects against apoptosis induced by calcium ionophore and amyloid beta-peptide.

The calcium binding protein, calbindin-D28k is normally present in neurons. Recently we reported that brain injury and tumor necrosis factors (TNFs) induce calbindin-D28k in astrocytes. TNF-treated calbindin expressing astrocytes were resistant to acidosis and calcium ionophore toxicity, suggesting that calbindin may have a cytoprotective role in astrocytes in the injured brain (M.P. Mattson, B. Cheng, S.A. Baldwin, V.L. Smith-Swintosky, J. Keller, J. Geddes, Scheff, J.W., Christakos, S., Brain injury and tumor necrosis factors induce calbindin-D28k in astrocytes: evidence for a cytoprotective response, J. Neurosci. Res., 42 (1995) 257). In order to obtain direct evidence for a role of calbindin, using the eukaryotic expression vector pREP4, rat calbindin-D28k was stably expressed in C6 rat astocytoma glial cells. Cytotoxicity in response to calcium ionophore or amyloid beta-peptide (which accumulates in the brain in Alzheimer's disease and has been reported to be neurotoxic) was measured by MTT reduction in vector transfected cells and in calbindin transfected clones. Stably expressed calbindin resulted in increased cell survival in the presence of calcium ionophore (1-10 microM) or amyloid beta-peptide (10-100 microM). In addition, the calcium ionophore or amyloid beta-peptide mediated rise in intracellular calcium in vector transfected cells was significantly attenuated in calbindin expressing cells. Apoptotic cell death was detected by the Hoechst method in vector transfected C6 glial cells treated with calcium ionophore or beta-amyloid (34-36% apoptotic cells/culture). The number of apoptotic nuclei was significantly attenuated in similarly treated calbindin-D28k transfected clones (10-13% apoptotic cells/culture; p<0.01). Our results support the involvement of calcium fluxes in apoptosis and suggest that calbindin-D28k, by buffering calcium, can suppress death in apoptosis susceptible cells in the central nervous system.

Amyloid beta-Peptides↗

PubMed1988

The 28-kDa calbindin-D is a major calcium-binding protein in the basilar papilla of the chick.

In previous work we identified a basilar papilla protein (BPP23) that appears to be one of the most abundant soluble proteins in the basilar papilla of the chick cochlea. Here we report the purification of protein BPP23 from chick cochlea and the generation of a specific antiserum. Immunoblotting and immunoprecipitation experiments with this antiserum indicate that BPP23 is a calcium-binding protein very similar, if not identical, to avian calbindin, the 28-kDa vitamin D-dependent calcium-binding protein. Although the basilar papilla contains both receptor hair cells and supporting cells, immunocytochemical studies by others have localized calbindin-like immunoreactivity to the hair cells in the rat auditory receptor epithelium. Our estimates of the abundance of protein BPP23, assuming exclusive localization within the hair cell, indicate a concentration of at least 1 mM. Avian calbindin has four high-affinity (Kd = 0.5 X 10(-6)) calcium-binding sites. The presence of a specific calcium-binding activity at such high levels suggests an important function for cochlear calbindin (BPP23) in hair cell calcium homeostasis and auditory transduction.

Animals↗

PubMed1990

Peptidyl-prolyl cis-trans isomerase does not affect the Pro-43 cis-trans isomerization rate in folded calbindin D9k.

The calcium-binding protein calbindin D9k has previously been shown to exist in two folded forms only differing in the proline cis-trans isomerism of the Gly-42-Pro-43 amide bond. This bond is located in a flexible loop connecting the two EF-hand Ca2+ sites. Calbindin D9k therefore constitutes a unique test case for investigating if the recently discovered enzyme peptidyl-prolyl cis-trans isomerase (PPIase) can affect the cis-trans exchange rate in a folded protein. The 1H NMR saturation transfer technique has been used to measure the rate of interconversion between the cis and trans forms of calbindin in the presence of PPIase (PPIase:calbindin concentration ratio 1:10) at 35 degrees C. No rate enhancement could be detected.

Amino Acid Isomerases↗

PubMed1988

Egg shell quality and cholecalciferol metabolism in aged laying hens.

Calcium-binding protein D28K (calbindin) synthesis, vitamin D metabolism and shell quality were investigated in young and aged laying hens fed diets containing either cholecalciferol (CC) or its 1-hydroxylated derivatives. Duodenal calbindin concentration was similar in the young and in the aged laying hens. Exogenous 1-hydroxylated CC derivatives increased duodenal calbindin concentration, regardless of age. Shell weight and shell density (mg/cm2) were significantly lower (P less than 0.01) in the aged than in the young hens. Egg shell weight and density tended to decrease along the clutch. The rate of decline was higher in aged than in young hens. Feeding aged hens a diet containing 5 micrograms 1,25-dihydroxycholecalciferol [1,25(OH)2CC] or 1 alpha-hydroxycholecalciferol per kilogram improved shell quality, slowed down the progressive reduction in shell quality during the clutch and increased culling and mortality. The results indicate a) that the capacity for expression of 1,25(OH)2D3 in the intestine is not altered by age and b) that prolonged feeding of 1-hydroxylated derivatives of vitamin D3 improves shell quality in aged laying hens and increases culling and mortality.

25-Hydroxyvitamin D3 1-alpha-Hydroxylase↗

PubMed1994

Identification of calbindin D-9k mRNA and its regulation by 1,25-dihydroxyvitamin D3 in Caco-2 cells.

We have identified and quantified specific mRNAs in the human colonic carcinoma cell line Caco-2 by reverse transcriptase-polymerase chain reaction. Initial examination revealed that like rat duodenal mucosa, Caco-2 cells possessed mRNA for the vitamin D receptor. Using primers for human calbindin we found a 237-bp PCR product in Caco-2 cell RNA, but not from rat duodenal RNA. Primers for rat calbindin did not amplify calbindin mRNA in Caco-2 RNA, confirming a high degree of mismatch between rat and human sequences. 1,25(OH)2 vitamin D3 treatment (10 nM) significantly elevated calbindin mRNA levels 50% by 12 h, with maximal levels occurring by 48 h (fivefold elevation). Increasing concentrations of 1,25(OH)2 vitamin D3 (from 15 pM to 100 nM) caused progressive increases in calbindin mRNA levels following 48 h of treatment. Elevated calbindin mRNA levels were associated with enhancement of transcellular calcium transport. Our results are the first demonstration of vitamin D-regulated calbindin mRNA in a human intestinal cell line.

Animals↗

PubMed1993

Calcium binding by chick calretinin and rat calbindin D28k synthesised in bacteria.

Calretinin is a member of the EF-hand calcium-binding protein family, with a high similarity with calbindin D28k. The chick calretinin cDNA sequence was reconstructed in a M13 vector and transferred into an expression plasmid derived from the pET series. The calretinin gene was expressed in Escherichia coli and produced immunoreactive calretinin of the expected size. Bacterially expressed calretinin was purified with successive ammonium-sulfate precipitation, DEAE chromatography, hydroxyapatite chromatography, Sephadex G-75 chromatography and Mono-Q chromatography. Normally, 1.0-1.5 mg calretinin was obtained from 1 l bacterial culture with a protein recovery of 0.5-1.5%. Calbindin D28k was purified similarly from bacteria using an expression plasmid provided by W. Hunziker. Calcium-binding activity of purified proteins was measured by equilibrium dialysis in calcium/EGTA mixtures with 45Ca as tracer. Both calretinin and calbindin D28k bound 3-4 Ca2+/molecule (calretinin, 4.0 +/- 0.5; calbindin D28k, 3.5 +/- 0.4), implying that at least one of the canonical EF-hand domains does not bind calcium. The Kd was 0.3-0.5 microM with little difference between the values for the two proteins.

Amino Acid Sequence↗

PubMed1992

Dietary calcium modifies concentrations of lead and other metals and renal calbindin in rats.

We studied the effects of dietary calcium on kidney, femur, testis, liver, heart and brain concentrations of lead, magnesium, iron, copper, calcium and zinc in rats exposed to lead for 1 y. Renal levels of the 28,000 Da, vitamin D-dependent, calcium-binding protein calbindin-D28K were also measured. Seventy-two weanling male Sprague-Dawley rats were randomly assigned to one of nine treatment groups. Rats were fed diets containing 0.1, 0.5 or 2.5% Ca for 52 wk and were simultaneously given either 0, 50 or 100 mg lead/L in their drinking water. Rats fed the 0.1% Ca diet had organ lead concentrations that were two- to 20-fold greater than the corresponding animals fed 0.5% Ca. Rats fed diets containing 2.5% Ca had the lowest organ lead concentrations. Despite substantial effects of diet Ca on organ lead concentrations, Ca did not significantly influence concentrations of most other divalent metals studied with the exception of kidney calcium and magnesium, testis iron, plasma ionic calcium and magnesium, and several femur metals. Kidney calcium concentrations were lower in rats fed 2.5% Ca diets than in those fed 0.1 or 0.5% Ca diets. For rats not given lead, renal calbindin concentrations were highest in rats fed 0.1% Ca, and lowest in rats fed 2.5% Ca. Lead inhibited an increase in renal calbindin in the rats fed 0.1% Ca, but paradoxically increased renal calbindin levels in animals fed 2.5% Ca.

Administration, Oral↗

PubMed2006

Influence of the "open field" exposure on calbindin D28K, calretinin, and parvalbumin containing cells in the rat midbrain - developmental study.

The aim of our study was to analyze the influence of the open field (OF) exposure on: 1. Distribution of c-Fos positive nuclei in: ventral tegmental area, substantia nigra, periaqueductal gray. 2. Appearance of calbindin-D28k, calretinin and parvalbumin in midbrain neurons that are engaged in the stress response. 3. Changes of c-Fos and calcium-binding proteins expression during maturation. The material consisted of Wistar rats of age between 0 and 90 days. The OF exposure was applied throughout 10 min and 90 min before the death of the animals. The brain sections were double stained using the antibodies against c-Fos, CB, CR or PV. Our results showed that in all studied nuclei age-related increase of c-Fos expression (without changing of its distribution properties) was found. PV didn't show any co-localization with c-Fos in neurons of studied regions at any ages, however some PV-immunoreactive (PV-ir) basket-like structures around c-Fos-immunoreactive (c-Fos-ir) neurons were observed. In the youngest group of rats c-Fos-ir cells and cells immunoreactive for CB and CR constituted separate neuronal populations. During maturation increases in the level of their co-localization with c-Fos was observed. We may conclude that in adult rat midbrain structures CB-immunoreactive (CB-ir) and CR-immunoreactive (CR-ir) cells (probably projection neurons) are mainly activated in the stress response following OF exposure. In the contrary PV-ir cells has only an indirect (modulatory) influence upon the c-Fos-ir cells.

Animals↗

PubMed2000

GABA(A) receptors on calbindin-immunoreactive myenteric neurons of guinea pig intestine.

These studies were carried out to characterize the properties of gamma-aminobutyric acidA (GABA(A)) receptors on guinea pig intestinal myenteric neurons maintained in primary culture. In addition, the type of neuron expressing GABA(A) receptors was identified using immunohistochemical methods. Whole-cell patch clamp recordings of currents elicited by GABA and acetylcholine (ACh) were obtained using pipettes containing Neurobiotin. After electrophysiological studies, neurons were processed for localization of calbindin-D28K-immunoreactivity (calbindin-ir). GABA (1 mM) and ACh (3 mM) caused inward currents in most cells tested. GABA currents were mimicked by muscimol (1-300 microM) and were blocked by bicuculline (10 microM) indicating that GABA was acting at GABA(A) receptors. GABA currents were associated with a conductance increase and a linear current/voltage relationship with a reversal potential of 1 +/- 1 mV (n = 5). Pentobarbital (PB, 3-1000 microM) and diazepam (DZP, 0.01-10 microM) potentiated GABA-induced currents. A maximum concentration of DZP (1 microM) increased GABA-induced currents 3.1 +/- 0.3 times while PB (1000 microM) increased GABA currents by 11 +/- 2 times. In outside-out patches, the amplitude of GABA-activated single-channel currents was linearly related to membrane potential with a single-channel conductance of 28.5 + 0.5 pS (n = 10). PB and DZP increased the open probability of GABA-induced single-channel currents. Neurons containing calbindin-ir were large, were isolated from other neurons and had GABA current amplitudes of -3.4 +/- 0.3 nA (n = 48). Neurons with weak or absent calbindin-ir were smaller, were localized in clusters of cells and had GABA-induced current amplitudes of -0.6 +/- 0.1 nA (n = 20). ACh-induced currents were smaller in calbindin-ir neurons (-0.7 +/- 0.1 nA) compared to weakly calbindin-ir neurons (-1.4 +/- 0.1 nA). These results indicate that myenteric calbindin-ir neurons express a high density of GABA(A) receptors. Cell size and location allow visual identification of neurons likely to contain calbindin-ir permitting targeted studies of the properties of these neurons.

Animals↗

PubMed1994

Identification of osteocalcin mRNA in nonosteoid tissue of rats and humans by reverse transcription-polymerase chain reaction.

Diseased or necrotic tissue can become calcified in a way that resembles bone. We examined soft tissues for the presence and regulation of the mRNA for the bone-associated protein, osteocalcin (OC). RNA was isolated from liver, kidney, lung, brain, muscle, and bone of young (2 months) male SD rats and analyzed for beta-actin, IGF-I, metallothionein IIa, alpha 1 collagen, calbindin-D9k (CaBP), and OC mRNA by reverse transcription-polymerase chain reaction (RT-PCR). All PCR products but CaBP were found in bone; CaBP was present only in duodenum, kidney, and lung. OC product was detected in all tissues; the identity of the PCR product was confirmed by sequencing. Bone OC mRNA levels were calculated to be 1000-fold higher than duodenal levels. Rats fed a 0.8% strontium diet for 7 days to drive down serum 1,25-dihydroxyvitamin D3 levels [1,25(OH)2D3] and then injected with 300 ng 1,25(OH)2D3/100 body weight had increased duodenal CaBP (2.5-fold) and femur OC mRNA (2.2-fold) 24 h after treatment. Duodenal OC mRNA was unchanged. OC mRNA was found in nondiseased human aortae, and the amount of message was elevated in calcified aorta and calcified aortic plaques. These results demonstrate that (1) tissues other than bone have low basal expression of OC mRNA, (2) OC mRNA is not regulated by vitamin D in nonosteoid tissue, and (3) expression of OC mRNA in atherosclerotic aorta reflects a role for bone-forming cells in ectopic bone formation observed in certain disease conditions.

Animals↗

PubMed2002

Molecular analysis of the vagal motoneuronal degeneration after right vagotomy.

The aim of this study was to investigate the vagal motoneuronal degeneration after right vagotomy using in situ hybridization, RT-PCR, and immunohistochemistry methods. The morphology of the vagal motoneurons in dorsal motor nucleus of the vagus nerve (DMV) and nucleus of ambiguus (NA) after right vagotomy was examined by using Nissl staing and TUNEL. The expression of inducible nitric oxide synthase (iNOS), bcl-2, bax, and caspase-3 in DMV and NA of rats after right vagotomy was studied. Additionally, the involvement of the N-methyl-D-aspartate (NMDA) receptor-calcium-neuronal nitric oxide synthase (nNOS) pathway in the vagal motoneuronal degeneration was addressed by double-immunolabeling analysis of nNOS with NMDAR1 and calbindin D28K in right-vagotomized rats. The neurons in right DMV and NA displayed a darkly stained, shrunken morphology at 1 day and 5 days following right vagotomy as shown by Nissl staining. Quantitative analysis revealed that, at 1 day and 5 days following right vagotomy, the number of neurons in right DMV, but not NA, was significantly reduced in comparison with that of control rats. Occasional TUNEL-positive neurons were detected in right DMV of rat at 1 day after right vagotomy. The expression of iNOS protein and mRNA was absent in DMV and NA of control rats. However, the iNOS mRNA expression was induced bilaterally in DMV and NA at 1 day postoperation and continued to be up-regulated until 5 days after vagotomy as shown by in situ hybridization. Immunohistochemistry analysis also showed the increased expression of iNOS in bilateral DMV and NA of vagotomized rats. RT-PCR analysis revealed the enhanced bcl-2 and reduced bax mRNA levels and subsequent up-regulation of both bcl-2 and bax mRNA in right sides of the vagotomized brainstems at 1 day and 5 days postoperation, respectively. In situ hybridization analysis confirmed the up-regulation of bcl-2 and bax mRNA in right DMV and NA of the rats at 5 days following operation. Immunohistochemistry analysis showed up-regulated Bcl-2 immunoreactivity and undetectable changes in Bax immunoreactivity in DMV and NA of rats at 1 day after vagotomy, whereas enhancement of both Bcl-2 and Bax immunoreactivity was observed at 5 days postoperation. In addition, the caspase-3 mRNA level was elevated ipsilaterally in DMV and NA at 1 day and 5 days following right vagotomy. Double-immunofluorescence analysis showed complete colocalization of nNOS with NMDAR1 and with calbindin in ipsilateral DMV and NA at 10 days following right vagotomy. This study suggests that the signal pathway for NMDAR1-calcium-nNOS and the up-regulation of iNOS in DMV and NA may be involved in the vagal motor neurodgeneration after right vagotomy. Furthermore, our results imply that the apoptosis pathway mediated by Bcl-2, Bax, and caspase-3 may be activated in vagal motoneurons after right vagotomy.

Animals↗

PubMed1998

Calbindin-D28k buffers intracellular calcium and promotes resistance to degeneration in PC12 cells.

The calcium-binding protein calbindin-D28k (CB) has been hypothesized to function, in part, as a neuroprotective protein. CB is localized within nerve cells that are often less vulnerable to degeneration in patients with Alzheimer's disease and Parkinson's disease, and cells containing CB can buffer intracellular calcium concentrations ([Ca2+]i). The present study was designed to directly test the hypothesis that CB can protect cells from degeneration by reducing [Ca2+]i. PC12 cells, transfected to express different levels of CB, were found to be significantly less vulnerable to degeneration caused by serum withdrawal, glutamate, and the neurotoxin 1-methyl-4-phenylpyridinium (MPP+). However, CB did not protect cells from degeneration caused by the calcium ionophore A23187. CB-transfected cells exhibited reduced elevations in [Ca2+]i following treatment with bradykinin, or ATP compared to non-CB-containing cells. These data indicate that CB can protect cells from degeneration caused by certain conditions, and it reduces elevations in [Ca2+]i caused by influx from extracellular sources.

1-Methyl-4-phenylpyridinium↗