Calbindin D28k Research Articles

Altered cerebellar organization and function in monoamine oxidase A hypomorphic mice

Monoamine oxidase A (MAO-A) is the key enzyme for the degradation of brain serotonin (5-hydroxytryptamine, 5-HT), norepinephrine (NE) and dopamine (DA). We recently generated and characterized a novel line of MAO-A hypormorphic mice (MAO-ANeo), featuring elevated monoamine levels, social deficits and perseverative behaviors as well as morphological changes in the basolateral amygdala and orbitofrontal cortex. Here we showed that MAO-ANeo mice displayed deficits in motor control, manifested as subtle disturbances in gait, motor coordination, and balance. Furthermore, magnetic resonance imaging of the cerebellum revealed morphological changes and a moderate reduction in the cerebellar size of MAO-ANeo mice compared to wild type (WT) mice. Histological and immunohistochemical analyses using calbindin-D-28k (CB) expression of Purkinje cells revealed abnormal cerebellar foliation with vermal hypoplasia and decreased in Purkinje cell count and their dendritic density in MAO-ANeo mice compared to WT. Our current findings suggest that congenitally low MAO-A activity leads to abnormal development of the cerebellum.

Overexpression of melatonin membrane receptors increases calcium‐binding proteins and protects VSC4.1 motoneurons from glutamate toxicity through multiple mechanisms

Melatonin has shown particular promise as a neuroprotective agent to prevent motoneuron death in animal models of both amyotrophic lateral sclerosis (ALS) and spinal cord injuries (SCI). However, an understanding of the roles of endogenous melatonin receptors including MT1, MT2, and orphan G-protein receptor 50 (GPR50) in neuroprotection is lacking. To address this deficiency, we utilized plasmids for transfection and overexpression of individual melatonin receptors in the ventral spinal cord 4.1 (VSC4.1) motoneuron cell line. Receptor-mediated cytoprotection following exposure to glutamate at a toxic level (25 μm) was determined by assessing cell viability, apoptosis, and intracellular free Ca(2+) levels. Our findings indicate a novel role for MT1 and MT2 for increasing expression of the calcium-binding proteins calbindin D28K and parvalbumin. Increased levels of calbindin D28K and parvalbumin in VSC4.1 cells overexpressing MT1 and MT2 were associated with cytoprotective effects including inhibition of proapoptotic signaling, downregulation of inflammatory factors, and expression of prosurvival markers. Interestingly, the neuroprotective effects conferred by overexpression of MT1 and/or MT2 were also associated with increases in the estrogen receptor β (ERβ): estrogen receptor α (ERα) ratio and upregulation of angiogenic factors. GPR50 did not exhibit cytoprotective effects. To further confirm the involvement of the melatonin receptors, we silenced both MT1 and MT2 in VSC4.1 cells using RNA interference technology. Knockdown of MT1 and MT2 led to an increase in glutamate toxicity, which was only partially reversed by melatonin treatment. Taken together, our findings suggest that the neuroprotection against glutamate toxicity exhibited by melatonin may depend on MT1 and MT2 but not GPR50.

Characterization of vitamin D-deficient klotho-/- mice: do increased levels of serum 1,25(OH)2D3 cause disturbed calcium and phosphate homeostasis in klotho-/- mice?

Klotho(-/-) mice display disturbed Ca(2+) and vitamin D homeostasis. Renal cytochrome p450 27b1 (Cyp27b1), the enzyme that catalyzes the hydrolysis to 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), is increased in klotho(-/-) mice, and a 1,25(OH)(2)D(3)-deficient diet partially normalized Ca(2+) homeostasis in these klotho(-/-) mice. The aim of the present study was to further delineate the interplay between 1,25(OH)(2)D(3) and klotho and their relative contribution to the Ca(2+) homeostasis of klotho(-/-) mice. Double-klotho(-/-)/Cyp27b1(-/-) mice were generated and mice aged 8-12 weeks were housed in metabolic cages to collect 24-h urine. Blood samples were taken and the animals were sacrificed, and the kidney and duodenum tissues were sampled for RNA extraction. The bone was fixed in 10% v/v formalin and analysed by microcomputed tomography (μCT) scans. Klotho(-/-)/Cyp27b1(-/-) mice, like Cyp27b1(-/-) mice, displayed significantly decreased serum total calcium concentrations compared with wild-type mice (1.44 ± 0.03 and 2.25 ± 0.02 mM) along with normal urinary total calcium excretion. Hyperphosphataemia of klotho(-/-) mice normalized to wild-type levels in klotho(-/-)/Cyp27b1(-/-) mice. The mRNA levels of duodenal transient receptor potential vanilloid subtype 6 (TRPV6) and calcium-binding protein-D(9K), and renal calbindin-D(28K) and NCX1 were significantly reduced in the double knockouts compared with wild-type or klotho(-/-) mice. Elevated TRPV5 protein levels in klotho(-/-) mice normalized to wild type in klotho(-/-)/Cyp27b1(-/-) mice, but were decreased in Cyp27b1(-/-) mice. μCT scans showed that klotho(-/-)/Cyp27b1(-/-) mice, as Cyp27b1(-/-) mice, display significant bone hypomineralization and severely decreased bone mass. Klotho(-/-) mice show a reduced bone mass and increased trabecular numbers. Klotho(-/-)/Cyp27b1(-/-) mice resemble Cyp27b1(-/-) mice. Since 1,25(OH)(2)D(3) is absent in these mice, our results imply that Ca(2+) homeostasis in klotho(-/-) mice is affected by their excessive 1,25(OH)(2)D(3) levels.

Two types of periglomerular cells in the olfactory bulb of the macaque monkey (Macaca fascicularis)

The olfactory bulb (OB) of mammals is the brain region that receives the sensory information coming from the olfactory epithelium. The entrance of the olfactory information occurs in spherical structures of neuropil named olfactory glomeruli and is modulated by a population of interneurons known as periglomerular cells (PG). It has been demonstrated that there are two types of PG in the OB of some macrosmatic mammals, including rats and mice. Type 1 PG (PG-1) receive synapses from the olfactory nerve, whereas type 2 PG (PG-2) do not receive synapses from the olfactory axons. To date, the presence of the two types of PG has not been investigated in microsmatic mammals. In this context, we analyze the presence of PG-1 and PG-2 in the OB of the long-tailed macaque (Macaca fascicularis). For that, we used the enzyme tyrosine hydroxylase, the neuronal isoform of the enzyme nitric oxide synthase and the calcium-binding proteins calbindin D-28k and calretinin as neurochemical markers. Our results demonstrate that the OB of the macaque contains PG-1 and PG-2. A subpopulation of PG-1 expresses tyrosine hydroxylase and another expresses the neuronal isoform of nitric oxide synthase. In addition, a subpopulation of PG-2 expresses calbindin D-28k and another expresses calretinin. Double immunofluorescence demonstrates that there is no colocalization of two markers in the same PG. These results mimic those found in macrosmatic animals. The presence of two types of PG in the glomerular circuits seems to be a key principle for the organization of the OB of mammals.

Expression of Calcium Transport Proteins in the Extraembryonic Membranes of a Viviparous Snake, virginia striatula

Yolk is the primary source of calcium for embryonic growth and development for most squamates, irrespective of mode of parity. The calcified eggshell is a secondary source for embryonic calcium in all oviparous eggs, but this structure is lost in viviparous lineages. Virginia striatula is a viviparous snake in which embryos obtain calcium from both yolk and placental transport of uterine calcium secretions. The developmental pattern of embryonic calcium acquisition in V. striatula is similar to that for oviparous snakes. Calbindin-D(28K) is a marker for epithelial calcium transport activity and plasma membrane Ca(2+)-ATPase (PMCA) provides the energy to catalyze the final step in calcium transport. Expression of calbindin-D(28K) and PMCA was measured by immunoblotting in yolk sac splanchnopleure and chorioallantois of a developmental series of V. striatula to test the hypothesis that these proteins mediate calcium transport to embryos. In addition, we compared the expression of calbindin-D(28K) in extraembryonic membranes of V. striatula throughout development to a previously published expression pattern in an oviparous snake to test the hypothesis that the ontogeny of calcium transport function is independent of reproductive mode. Expression of calbindin-D(28K) increased in yolk sac splanchnopleure and chorioallantois coincident with calcium mobilization from yolk and uterine sources and with embryonic growth. The amount of PMCA in the chorioallantois did not change through development suggesting its expression is not rate limiting for calcium transport. The pattern of expression of calbindin-D(28K) and PMCA confirms our initial hypothesis that these proteins mediate embryonic calcium uptake. In addition, the developmental pattern of calbindin-D(28K) expression in V. striatula is similar to that of an oviparous snake, which suggests that calcium transport mechanisms and their regulation are independent of reproductive mode.

Retinal horizontal cells reduced in a rat model of congenital stationary night blindness

This work was conducted to determine whether congenital stationary night blindness (CSNB), which is caused by a Cacna1f mutation, could affect development of second-order neurons in the retina, such as horizontal cells (HCs). The CSNB rats and age-matched wild type rats were sacrificed at postnatal days (PND) 15, 30 and 60. Morphometric analyses of HCs, which were labeled by a primary antibody to calbindin D-28K, were performed at the light microscopic level on retinal cross sections and whole mount retinas. Calbindin D-28K was measured by western blotting in retinal samples. We found that the average number and density of HCs, Calbindin level and thickness of OPL were all decreased significantly in CSNB group compared to control group. These results indicated that second-order retinal neurons, such as horizontal cells, are affected by retinal degeneration. The relationship between the absence of HCs and the gene defect of CSNB requires further research.

Sodium deoxycholate inhibits chick duodenal calcium absorption through oxidative stress and apoptosis

High concentrations of sodium deoxycholate (NaDOC) produce toxic effects. This study explores the effect of a single high concentration of NaDOC on the intestinal Ca2+ absorption and the underlying mechanisms. Chicks were divided into two groups: 1) controls and 2) treated with different concentrations of NaDOC in the duodenal loop for variable times. Intestinal Ca2+ absorption was measured as well as the gene and protein expressions of molecules involved in the Ca2+ transcellular pathway. NaDOC inhibited the intestinal Ca2+ absorption, which was concentration dependent. Ca2+-ATPase mRNA decreased by the bile salt and the same occurred with the protein expression of Ca2+-ATPase, calbindin D28k and Na+/Ca2+ exchanger. NaDOC produced oxidative stress as judged by ROS generation, mitochondrial swelling and glutathione depletion. Furthermore, the antioxidant quercetin blocked the inhibitory effect of NaDOC on the intestinal Ca2+ absorption. Apoptosis was also triggered by the bile salt, as indicated by the TUNEL staining and the cytochrome c release from the mitochondria. As a compensatory mechanism, enzyme activities of the antioxidant system were all increased. In conclusion, a single high concentration of NaDOC inhibits intestinal Ca2+ absorption through downregulation of proteins involved in the transcellular pathway, as a consequence of oxidative stress and mitochondria mediated apoptosis.

Immunohistochemical study of the distribution of calcium binding proteins in the brain of a chondrostean (Acipenser baeri)

Chondrosteans represent an ancient lineage in ray-finned bony fishes and hence in jawed vertebrates. This immunohistochemical study in the brain of the Siberian sturgeon reports the neuronal distribution of three cytosolic calcium-binding proteins: calbindin-D28k (CB), calretinin (CR), and parvalbumin (PV). CB and CR are widely expressed in different neuron subsets distributed throughout the sturgeon brain. Studies using double immunofluorescence reveal a wide co-distribution of CB and CR in the brain nuclei but scarce co-localization at cellular level. In the forebrain, CR- and CB-immunoreactive (ir) populations were observed in the olfactory bulbs, in pallial and subpallial telencephalic areas, and in some diencephalic nuclei. CR-ir cells were also observed in the posterior tubercle and CB-ir cells in the preglomerular complex. At midbrain and hindbrain basal levels, CB-ir and CR-ir cell bodies were mainly distributed in periventricular areas. In the cerebellum, CB and CR cells were co-localized in some granular cell subsets in laterodorsal and dorsolateral regions, and in some Purkinje-like cells. CB-ir and CR-ir fibers were mainly observed in the olfactory bulbs, hypothalamus, and habenula, and in fiber tracts that coursed in the optic tectum and through the mesencephalic and rhombencephalic basal areas. With regard to PV, the sturgeon brain showed a rather limited distribution of PV-ir perikarya and fibers. Thus, CR, CB, and PV allowed the identification of subpopulations of neurons not distinguished on the basis of cytoarchitecture alone, which provided a better understanding of the anatomical organization of the sturgeon brain. These results reveal numerous shared features with teleosts, but also important differences.

Decreased hippocampal expression of calbindin D28K and cognitive impairment in MELAS

Mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS) is a maternally inherited mitochondrial syndrome characterized by seizures, migrainous headaches, lactic acidosis, vomiting, and recurrent stroke-like episodes. Patients often suffer from cognitive dysfunction of unclear pathogenesis. In this study, we explored a possible link between cognitive dysfunction and hippocampal expression of calbindin D28KD (CB), a high affinity calcium-binding protein, in four MELAS patients, using post mortem hippocampal tissues. We found significantly reduced CB levels in all patients by immunohistochemistry, Western blot, and quantitative real-time PCR. Reduction in CB expression has been associated with aging and with neurodegenerative disorders, including Alzheimer's disease. We postulate that the reduced CB expression may play a role in the cognitive abnormalities associated with MELAS.

Low oral doses of bisphenol A increase volume of the sexually dimorphic nucleus of the preoptic area in male, but not female, rats at postnatal day 21

Perinatal treatment with relatively high doses of bisphenol A (BPA) appears to have little effect on volume of the rodent sexually dimorphic nucleus of the preoptic area (SDN-POA). However, doses more relevant to human exposures have not been examined. Here, effects of pre- and post-natal treatment with low BPA doses on SDN-POA volume of postnatal day (PND) 21 Sprague–Dawley rats were evaluated. Pregnant rats were orally gavaged with vehicle, 2.5 or 25.0μg/kg BPA, or 5.0 or 10.0μg/kg ethinyl estradiol (EE2) on gestational days 6–21. Beginning on the day after birth, offspring were orally treated with the same dose their dam had received. On PND 21, offspring (n=10–15/sex/group; 1/sex/litter) were perfused and volume evaluation was conducted blind to treatment. SDN-POA outline was delineated using calbindin D28K immunoreactivity. Pairwise comparisons of the significant treatment by sex interaction indicated that neither BPA dose affected female volume. However, females treated with 5.0 or 10.0μg/kg EE2 exhibited volumes that were larger than same-sex controls, respectively (p<0.001). Males treated with either BPA dose or 10.0μg/kg/day EE2 had larger volumes than same-sex controls (p<0.006). These data indicate that BPA can have sex-specific effects on SDN-POA volume and that these effects manifest as larger volumes in males. Sensitivity of the methodology as well as the treatment paradigm was confirmed by the expected EE2-induced increase in female volume. These treatment effects might lead to organizational changes within sexually dimorphic neuroendocrine pathways which, if persistent, could theoretically alter adult reproductive physiology and socio-sexual behavior in rats.

The changes of calretinin immunoreactivity in paraquat-induced nephrotoxic rats

Calcium-binding proteins are present in the kidneys: calbindin D-28k in the distal tubules and calretinin in the proximal tubules. Since paraquat causes degeneration in the brush border-bearing proximal tubule cells in rat kidneys, we investigated the changes of calretinin immunoreactivity in the proximal tubule cells of paraquat-induced nephrotoxicity in experimental male Sprague-Dawley rats following chitosan oligosaccharide pretreatment to investigate its protective properties. Paraquat (60mg/kg) was administered intraperitoneally with or without chitosan oligosaccharide (500mg/kg, p.o.) pretreatment. The changes on calretinin were compared with those of calbindin D-28k by immunohistochemistry and Western Blot analysis. Calretinin was immunolocalized on the apical surface of proximal tubule cells in the deeper cortex of normal kidney, and disappeared after paraquat administration with minor changes of calbindin D-28k immunoreactivity in the distal tubules and collecting ducts. Chitosan oligosaccharide pretreatment caused increased expression of calretinin and calbindin D-28k before paraquat injection and helped preserve proximal tubules after paraquat treatment. However, Western blot analysis on calretinin and calbindin D-28k could not explain the degeneration of the proximal tubule cells in paraquat-induced nephrotoxicity. These findings suggested that calretinin is a possible and more useful histopathological marker for proximal tubule cells in paraquat-induced nephrotoxic rats.

The Decline of Autophagy Contributes to Proximal Tubular Dysfunction During Sepsis

Severe sepsis associated with overproduction of tumor necrosis factor α and reactive oxygen species leads to energy depletion and cellular damage. Both reactive oxygen species and damaged organelles induce autophagy for recycling nutrients to combat pathological stress. To study whether autophagy plays a beneficial role in the pathogenesis of renal failure during sepsis, rats were subjected to cecal ligation and puncture (CLP) or sham operation. Temporal relationship of autophagy and renal dysfunction were examined in vivo. The results showed that the level of lipidated microtubule-associated protein light chain 3 (LC3-II), a marker of autophagy, elevated transiently at 3 h but declined at 9 h until 18 h after CLP. Light chain 3 aggregation in renal tissue showed a similar trend to the change of LC3-II protein. High levels of blood urea nitrogen and creatinine as well as low tubular sodium reabsorption occurred at 18 h after CLP. The distribution of autophagy located primarily in angiotensin-converting enzyme-positive, which is concentrated in proximal tubule, but calbindin D28k (calcium-binding protein D28K, a marker of distal tubule)-negative cells in renal cortex. Therefore, NRK-52E (proximal tubule epithelial cell line) cells were used to further examine cell viability and DNA fragmentation after silencing or inducing autophagy. We found that knockdown of Atg7 (autophagy-related gene 7) exaggerates, whereas preincubation of rapamycin (an autophagy inducer) diminishes tumor necrosis factor α-induced cell death. These results suggest that the decline of sepsis-induced autophagy contributes to the proximal tubular dysfunction, and maintenance of sufficient autophagy prevents cell death. These data open prospects for therapies that activate autophagy during sepsis.

Differential alterations of synaptic plasticity in dentate gyrus and CA1 hippocampal area of Calbindin-D28K knockout mice

Regulation of the intracellular calcium concentration ([Ca2+]i) is of critical importance for synaptic function. Therefore, neurons buffer [Ca2+]i using intracellular Ca2+-binding proteins (CaBPs). Previous evidence suggests that Calbindin-D28K (CB), an abundantly expressed endogenous fast CaBP, plays an important role in neuronal survival, motor coordination, spatial learning paradigms and some forms of synaptic plasticity. In the present study, the role of CB in synaptic transmission and plasticity was further investigated using extracellular recordings of synaptic activity in cell- and dendritic layers of dentate gyrus (DG) and CA1 area in hippocampal slices from wild-type, heterozygous and homozygous CB knockout mice. The results demonstrate a consistent failure to maintain long-term potentiation (LTP) in hippocampal DG and CA1 area of knockout mice. Compared to wild-type mice, the paired-pulse ratio of EPSPs recorded in DG is significantly lower in slices from knockout mice, whereas it is significantly higher in CA1 area. The amplitude of the population spike recorded in CA1 area of wild-type mice steadily increases following tetanic stimulation, whereas it steadily decreases in knockout mice. The combined results demonstrate that the absence of CB results in an impairment of LTP maintenance in both hippocampal DG and CA1 area, whereas paired-pulse facilitation and cellular excitability in CA1 area are differentially affected. These results support the role of CB as a critical determinant for several forms of synaptic plasticity in hippocampal DG and CA1 area. It is hypothesized that CB functions as a postsynaptic Ca2+ buffer as well as a presynaptic Ca2+ sensor.

Effects of in utero endotoxemia on the ovine fetal brain A model for schizophrenia

Infections during pregnancy can adversely affect the development of the fetal brain. This may contribute to disease processes such as schizophrenia in later life. Changes in the (cyto-) architecture of the anterior cingulate cortex (ACC), particularly in GABA-ergic interneurons, play a role in the pathogenesis of schizophrenia. We hypothesized that exposure to infection during pregnancy could result in cyto-architectural changes in the fetal ACC, similar to the pathogenesis seen in schizophrenia. Fetal sheep of 110 days GA (term=150 days GA) received an intravenous injection of 100 ng or 500 ng lipopolysaccharide (LPS) or saline as control. After delivery at 113 days GA, the cyto-architecture of the cingulate cortex (CC) was examined by immunohistochemistry. High dose LPS exposure resulted in a decreased density of GFAP-, calbindin D-28K- and parvalbumin-immunoreactive cells in the CC. In addition, these cells and calretinin-immunoreactive cells showed a changed morphology with reduced cell processes. This study provides further evidence that intra-uterine endotoxemia can induce changes in the fetal brain which correspond with changes seen in schizophrenia.

Absence of the calcium-binding protein calretinin, not of calbindin D-28k, causes a permanent impairment of murine adult hippocampal neurogenesis

Calretinin (CR) and calbindin D-28k (CB) are cytosolic EF-hand Ca2+-binding proteins and function as Ca2+ buffers affecting the spatiotemporal aspects of Ca2+ transients and possibly also as Ca2+ sensors modulating signaling cascades. In the adult hippocampal circuitry, CR and CB are expressed in specific principal neurons and subsets of interneurons. In addition, CR is transiently expressed within the neurogenic dentate gyrus (DG) niche. CR and CB expression during adult neurogenesis mark critical transition stages, onset of differentiation for CR, and the switch to adult-like connectivity for CB. Absence of either protein during these stages in null-mutant mice may have functional consequences and contribute to some aspects of the identified phenotypes. We report the impact of CR- and CB-deficiency on the proliferation and differentiation of progenitor cells within the subgranular zone (SGZ) neurogenic niche of the DG. Effects were evaluated (1) two and four weeks postnatally, during the transition period of the proliferative matrix to the adult state, and (2) in adult animals (3 months) to trace possible permanent changes in adult neurogenesis. The absence of CB from differentiated DG granule cells has no retrograde effect on the proliferative activity of progenitor cells, nor affects survival or migration/differentiation of newborn neurons in the adult DG including the SGZ. On the contrary, lack of CR from immature early postmitotic granule cells causes an early loss in proliferative capacity of the SGZ that is maintained into adult age, when it has a further impact on the migration/survival of newborn granule cells. The transient CR expression at the onset of adult neurogenesis differentiation may thus have two functions: (1) to serve as a self-maintenance signal for the pool of cells at the same stage of neurogenesis contributing to their survival/differentiation, and (2) it may contribute to retrograde signaling required for maintenance of the progenitor pool.

Three functional facets of calbindin D-28k

Many neurons of the vertebrate central nervous system (CNS) express the Ca2+ binding protein calbindin D-28k (CB), including important projection neurons like cerebellar Purkinje cells but also neocortical interneurons. CB has moderate cytoplasmic mobility and comprises at least four EF-hands that function in Ca2+ binding with rapid to intermediate kinetics and affinity. Classically it was viewed as a pure Ca2+ buffer important for neuronal survival. This view was extended by showing that CB is a critical determinant in the control of synaptic Ca2+ dynamics, presumably with strong impact on plasticity and information processing. Already 30 years ago, in vitro studies suggested that CB could have an additional Ca2+ sensor function, like its prominent acquaintance calmodulin (CaM). More recent work substantiated this hypothesis, revealing direct CB interactions with several target proteins. Different from a classical sensor, however, CB appears to interact with its targets both, in its Ca2+-loaded and Ca2+-free forms. Finally, CB has been shown to be involved in buffered transport of Ca2+, in neurons but also in kidney. Thus, CB serves a threefold function as buffer, transporter and likely as a non-canonical sensor.

Secretagogin is Expressed in Sensory CGRP Neurons and in Spinal Cord of Mouse and Complements other Calcium-Binding Proteins, with a Note on Rat and Human

BackgroundSecretagogin (Scgn), a member of the EF-hand calcium-binding protein (CaBP) superfamily, has recently been found in subsets of developing and adult neurons. Here, we have analyzed the expression of Scgn in dorsal root ganglia (DRGs) and trigeminal ganglia (TGs), and in spinal cord of mouse at the mRNA and protein levels, and in comparison to the well-known CaBPs, calbindin D-28k, parvalbumin and calretinin. Rat DRGs, TGs and spinal cord, as well as human DRGs and spinal cord were used to reveal phylogenetic variations.ResultsWe found Scgn mRNA expressed in mouse and human DRGs and in mouse ventral spinal cord. Our immunohistochemical data showed a complementary distribution of Scgn and the three CaBPs in mouse DRG neurons and spinal cord. Scgn was expressed in ~7% of all mouse DRG neuron profiles, mainly small ones and almost exclusively co-localized with calcitonin gene-related peptide (CGRP). This co-localization was also seen in human, but not in rat DRGs. Scgn could be detected in the mouse sciatic nerve and accumulated proximal to its constriction. In mouse spinal cord, Scgn-positive neuronal cell bodies and fibers were found in gray matter, especially in the dorsal horn, with particularly high concentrations of fibers in the superficial laminae, as well as in cell bodies in inner lamina II and in some other laminae. A dense Scgn-positive fiber network and some small cell bodies were also found in the superficial dorsal horn of humans. In the ventral horn, a small number of neurons were Scgn-positive in mouse but not rat, confirming mRNA distribution. Both in mouse and rat, a subset of TG neurons contained Scgn. Dorsal rhizotomy strongly reduced Scgn fiber staining in the dorsal horn. Peripheral axotomy did not clearly affect Scgn expression in DRGs, dorsal horn or ventral horn neurons in mouse.ConclusionsScgn is a CaBP expressed in a subpopulation of nociceptive DRG neurons and their processes in the dorsal horn of mouse, human and rat, the former two co-expressing CGRP, as well as in dorsal horn neurons in all three species. Functional implications of these findings include the cellular refinement of sensory information, in particular during the processing of pain.

The visinin-like proteins VILIP-1 and VILIP-3 in Alzheimer's disease—old wine in new bottles

The neuronal Ca2+-sensor (NCS) proteins VILIP-1 and VILIP-3 have been implicated in the etiology of Alzheimer's disease (AD). Genome-wide association studies (GWAS) show association of genetic variants of VILIP-1 (VSNL1) and VILIP-3 (HPCAL1) with AD+P (+psychosis) and late onset AD (LOAD), respectively. In AD brains the expression of VILIP-1 and VILIP-3 protein and mRNA is down-regulated in cortical and limbic areas. In the hippocampus, for instance, reduced VILIP-1 mRNA levels correlate with the content of neurofibrillary tangles (NFT) and amyloid plaques, the pathological characteristics of AD, and with the mini mental state exam (MMSE), a test for cognitive impairment. More recently, VILIP-1 was evaluated as a cerebrospinal fluid (CSF) biomarker and a prognostic marker for cognitive decline in AD. In CSF increased VILIP-1 levels correlate with levels of Aβ, tau, ApoE4, and reduced MMSE scores. These findings tie in with previous results showing that VILIP-1 is involved in pathological mechanisms of altered Ca2+-homeostasis leading to neuronal loss. In PC12 cells, depending on co-expression with the neuroprotective Ca2+-buffer calbindin D28K, VILIP-1 enhanced tau phosphorylation and cell death. On the other hand, VILIP-1 affects processes, such as cyclic nucleotide signaling and dendritic growth, as well as nicotinergic modulation of neuronal network activity, both of which regulate synaptic plasticity and cognition. Similar to VILIP-1, its interaction partner α4β2 nicotinic acetylcholine receptor (nAChR) is severely reduced in AD, causing severe cognitive deficits. Comparatively little is known about VILIP-3, but its interaction with cytochrome b5, which is part of an antioxidative system impaired in AD, hint toward a role in neuroprotection. A current hypothesis is that the reduced expression of visinin-like protein (VSNLs) in AD is caused by selective vulnerability of subpopulations of neurons, leading to the death of these VILIP-1-expressing neurons, explaining its increased CSF levels. While the Ca2+-sensor appears to be a good biomarker for the detrimental effects of Aβ in AD, its early, possibly Aβ-induced, down-regulation of expression may additionally attenuate neuronal signal pathways regulating the functions of dendrites and neuroplasticity, and as a consequence, this may contribute to cognitive decline in early AD.

Cytarabine induced cerebellar neuronal damage in juvenile rat: Correlating neurobehavioral performance with cellular and genetic alterations

Cytosine arabinoside (Ara-C), a pyrimidine analogue induces cerebellar dysfunction and behavioral abnormalities. Although many in vitro experiments have been conducted in the past demonstrating the lethal potential of Ara-C to cerebellar neurons, there is a paucity of literature available regarding the effects of Ara-C on the cellular and genetic material of cerebellum and its subsequent influence on the neurobehavioral performance in vivo. Rats were treated with Ara-C at the dose levels 50, 100 and 200mg/kg/day for 5 and 14 days by intraperitoneal (i.p.) route. Endpoints of the evaluation included food and water intake, body and organ weight, behavioral parameters, histopathology, oxidative stress, DNA damage, apoptosis, expression of p53, caspase-3 and calbindin D-28K (calbindin) as well as histone acetylation and methylation. Ara-C treatment for 14 days significantly decreased the food and water intake, body weight gain and brain weight in rat as compared to the control. Alterations in various behavioral parameters were observed, indicating the impaired cerebellar function. Further, cellular abnormalities in the cerebellum such as Purkinje cell misalignment and granule cell cytotoxicity were observed. Positive correlation was observed between Ara-C induced disturbance in the motor performance and the Purkinje cell loss in rat cerebellum. Moreover, Ara-C treatment significantly increased the oxidative stress, DNA damage, TUNEL positive cells, p53 and caspase-3 positive cells in the rat cerebellum. Unlike short-term treatment, long-term Ara-C treatment significantly reduced calbindin expression in the cerebellum. Apart from this, 14 days Ara-C treatment led to significant alterations in the histone acetylation and methylation in the cerebellum, while in 5 days treatment no such alterations were observed. Present results indicated that Ara-C, by inducing oxidative stress mediated DNA damage, executes neuronal apoptosis which is accompanied by an increase in the p53 and caspase-3, but decrease in the calbindin expression.

Proliferative enteropathy (PE)-induced changes in the calbindin-immunoreactive (CB-IR) neurons of inferior mesenteric ganglion supplying the descending colon in the pig.

A subpopulation of the pig inferior mesenteric ganglia (IMG) neurons projecting to the colon exhibit calbindin-like immunoreactivity. It is not known if there are any changes in the chemical coding patterns of these neurons during porcine proliferative enteropathy (PE). To answer this question, juvenile Large White Polish pigs with clinically diagnosed Lawsonia intracellularis infection (PE; n = 3) and a group of uninfected controls (C; n = 3) were compared. The retrograde tracer fast blue (FB) was injected into the descending colons of all animals and then tissue comprising IMGs from both groups was processed for double-labeling immunofluorescence with calbindin-D28k (CB) in combination with either tyrosine hydroxylase (TH), neuropeptide Y (NPY), somatostatin (SOM), vasoactive intestinal polypeptide (VIP), nitric oxide synthase, Leu-enkephalin, substance P, vesicular acetylcholine transporter, galanin, or pituitary adenylate cyclase-activating polypeptide. Immunohistochemistry revealed changes in the chemical coding pattern of calbindin-immunoreactive neurons in the inferior mesenteric ganglia of the pig. In control animals, FB/CB-positive neurons were immunoreactive to TH, NPY, SOM, and VIP. In the experimental group, TH-expressing neurons were unaffected, NPY-expressing neurons were increased, whereas the number of neurons immunoreactive to SOM or VIP was reduced. Changes in chemical coding of CB neurons during PE may play an important role in adaptation of these IMG cells under pathological conditions.