Tissue- and cell-specific distribution of creatine kinase B: A new and highly specific monoclonal antibody for use in immunohistochemistry

Sistermans, Erik A.; Kok, Y.J.M. de; Peters, W.H.M.; Ginsel, L.A.; Jap, P. H. K.; Wieringa, B.

doi:10.1007/bf00307817

Cited by 57 publications

(51 citation statements)

References 52 publications

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“…The immunoblot in Fig. 1A confirms that uMtCK and CK-B are present in WT mouse retinas (12,13). The blot also shows that CK-B is absent in retinas from CK-B-/-mice (14) and that both isoforms are missing in retinas from CK-B-/-;uMtCK-/-("CK-/-dko") mice (15).…”

Section: Resultssupporting

confidence: 57%

“…Immunoelectron microscopy provided additional evidence that CK-B is absent from the OS (Fig. S2E) (12). There is little difference in the distribution of CK-B in light vs. dark (Fig.…”

Section: Resultsmentioning

confidence: 87%

“…A previous study detected CK-B in isolated bovine rod OS (13), but other studies of mouse (12) and chicken (31) retinas found CK-B only in photoreceptor inner segments and terminals. We used both immunological methods and biochemical activity measurements to confirm that CK is sequestered in the synaptic terminals of photoreceptors and absent from the OSs.…”

Section: Discussionmentioning

confidence: 85%

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Flow of energy in the outer retina in darkness and in light

Linton

Holzhausen

Babai

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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Structural features of neurons create challenges for effective production and distribution of essential metabolic energy. We investigated how metabolic energy is distributed between cellular compartments in photoreceptors. In avascular retinas, aerobic production of energy occurs only in mitochondria that are located centrally within the photoreceptor. Our findings indicate that metabolic energy flows from these central mitochondria as phosphocreatine toward the photoreceptor's synaptic terminal in darkness. In light, it flows in the opposite direction as ATP toward the outer segment. Consistent with this model, inhibition of creatine kinase in avascular retinas blocks synaptic transmission without influencing outer segment activity. Our findings also reveal how vascularization of neuronal tissue can influence the strategies neurons use for energy management. In vascularized retinas, mitochondria in the synaptic terminals of photoreceptors make neurotransmission less dependent on creatine kinase. Thus, vasculature of the tissue and the intracellular distribution of mitochondria can play key roles in setting the strategy for energy distribution in neurons.energy metabolism | phototransduction A significant energy distribution problem can arise from the relative locations of mitochondria, ion pumps, and synapses in neurons. In photoreceptors, ion pumps occupy the intervening space between the centrally located mitochondria and the synaptic terminal. Ion pumping in dark-adapted photoreceptors consumes ∼20× more energy than neurotransmission (1). Therefore, the pumps could intercept all the metabolic energy made by the mitochondria before it can reach the synaptic terminal. In the vascularized retinas of mice, rats, and humans (2-4) this problem is solved by the presence of additional mitochondria in the terminal. However, in the avascular retinas of zebrafish, salamanders, rabbits, and guinea pigs there are no mitochondria in the terminals (2, 4, 5), which creates a need to partition some of the energy made by the central mitochondria into a protected form that can bypass the ion pumps to support the essential energy demands of the synaptic terminal.Energy consumption within retinal photoreceptors is compartmentalized and light-dependent. During illumination, phototransduction and light adaptation consume energy in the outer segment (OS). In darkness, energy is consumed by ion pumps in the inner segment and by glutamate release at the synaptic terminal (1). Energy demands and O 2 consumption are far greater in darkness than in light (1, 6-8).Metabolic energy is distributed in most cells as either ATP or phosphocreatine (PCr). There are 2 isoforms of creatine kinase (CK) in neurons, ubiquitous mitochondrial creatine kinase (uMtCK), and brain-type cytoplasmic creatine kinase (CK-B). uMtCK creates PCr from ATP at mitochondria (9), and CK-B can recreate ATP from PCr at sites of energy demand. In this way uMtCK and CK-B can collaborate to transfer metabolic energy between neuronal compartments (10, 11). This paper descr...

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Section: Resultssupporting

confidence: 57%

“…Immunoelectron microscopy provided additional evidence that CK-B is absent from the OS (Fig. S2E) (12). There is little difference in the distribution of CK-B in light vs. dark (Fig.…”

Section: Resultsmentioning

confidence: 87%

Section: Discussionmentioning

confidence: 85%

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Flow of energy in the outer retina in darkness and in light

Linton

Holzhausen

Babai

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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“…The photoreceptor inner segment, where most of the mitochondria are located for oxidative phosphorylation in the retina, is highly metabolically active to produce energy and nucleotides for phototransduction in the outer segment. 12,13 There is evidence that the photoreceptor outer segment also is involved in anaerobic glycolysis and contributes to retinal energy production. 14 It has been reported that impairment of photoreceptor energy metabolism contributes to photoreceptor degeneration in several retinal diseases.…”

mentioning

confidence: 99%

Laser Capture Microdissection-Directed Profiling of Glycolytic and mTOR Pathways in Areas of Selectively Ablated Müller Cells in the Murine Retina

Chung¹,

Shen²,

Gillies³

2013

Invest. Ophthalmol. Vis. Sci.

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Citation: Chung SH, Shen W, Gillies MC. Laser capture microdissectiondirected profiling of glycolytic and mTOR pathways in areas of selectively ablated müller cells in the murine retina. Invest Ophthalmol Vis Sci. 2013;54:6578-6584. DOI:10.1167/ iovs.13-12311 PURPOSE. We have reported previously down-regulation of key metabolic pathways, the glycolytic and mTOR pathways, from a global retinal microarray analysis after selective Müller cell ablation in a novel transgenic model. The purpose of the present study was to examine changes in expression of key molecules of glycolytic and mTOR pathways specifically in patches of Müller cell loss.METHODS. Eyes were enucleated 1 and 3 months after induced Müller cell ablation, directly embedded in optimal cutting temperature medium, and snap frozen in liquid nitrogen. Laser capture microdissection (LCM) was conducted to dissect patches of Müller cell loss for quantitative RT-PCR (qRT-PCR) analysis of key genes of the glycolytic (glyceraldehyde-3-phosphate dehydrogenase, enolase 1 and 2, lactate dehydrogenase A and B) and mTOR pathways (insulin-like growth factor receptor 1, phosphatidylinositide-3-kinase, Akt1, and regulatory-associated protein of mTOR). Protein validations were performed by immunohistochemistry. RESULTS. The LCM-directed qRT-PCR analysis ofMüller cell ablated specimens demonstrated reduced transcription of genes involved in the glycolytic and mTOR metabolic pathways. Of the proteins we chose to study, only enolase 1 was expressed by Müller cells. Other glycolytic and mTOR pathway proteins were expressed by photoreceptor inner and outer segments, which were lost in patches of Müller cell ablation.CONCLUSIONS. We found suppression of genes encoding various glycolytic and mTOR pathwayassociated enzymes in areas of Müller cell loss. This appeared mainly to be due to loss of photoreceptor inner and outer segments. The consequences of metabolic derangement caused by Müller cell ablation warrant further investigation.Keywords: Müller cells, glycolysis, mTOR pathway, photoreceptors, laser capture microdissection M¨u ller cells are the principal glial cells of the mammalian retina. They provide an anatomic and functional link between retinal neurons and their surrounding compartments, since they span the entire thickness of the retina from the microvilli projecting from the outer limiting membrane, formed by the apical processes of Müller cells, to the Müller cell end feet, which form the inner limiting membrane. They have critical roles in maintaining retinal homeostasis, such as producing neuroactive substances through a neurotransmitter recycling system, 1 maintaining the blood-retinal barrier (BRB), 2 and regulating ion, water content, and pH. 3,4 Müller cells also provide metabolic support and nutrients to neurones via metabolic pathways, such as the glycolytic pathway. 5 There is strong evidence that the Müller cell is one of the major cells responsible for metabolizing glucose into pyruvate and lactate to fuel neurons in the retina. [6][7][8][9][10][11...

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“…A CKB-specific monoclonal antibody (mAb) CK-BYK/21E10, which recognizes residues 4-20 at the amino terminus of CKB [21], was incubated with the membrane overnight at 4°C as described [17]. In separate experiments, a rabbit polyclonal CKB-specific antibody raised against amino acid residues 258-270 of CKB [1] was used at a dilution of 1:5,000.…”

Section: Immunoblot Analysismentioning

confidence: 99%

Expression of Creatine Kinase Isoenzyme Genes during Postnatal Development of Rat Brain Cerebrum: Evidence for Posttranscriptional Regulation

Shen¹,

Willis²,

Zhang³

et al. 2003

Dev Neurosci

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Brain creatine kinase (CKB) has a central role in the regeneration of ATP in the brain. During postnatal development of rat brain cerebrum, the CKB protein level was very low at postnatal day 1 and week 1 but by week 4 had increased 6- to 7-fold and remained constant through week 10. Surprisingly, CKB mRNA levels were already maximal at postnatal day 1 and week 1, indicating that CKB protein expression does not simply reflect the levels of CKB mRNA and is likely regulated posttranscriptionally during early postnatal times. Interestingly, the majority of cytoplasmic CKB mRNA was found to be associated with polyribosomes both at postnatal day 3 and week 6. Therefore, low CKB protein levels at early postnatal times could either be due to (1) normal translation initiation of CKB mRNA followed by a subsequent arrest during elongation or termination and/or (2) normal translation of CKB mRNA followed by rapid degradation of CKB protein. However, CKB protein increased coincidently with ubiquitous mitochondrial CK protein, suggesting that a functional phosphocreatine energy shuttle is formed in the cerebrum during postnatal development. The apparent posttranscriptional regulation of CKB in early postnatal cerebrum contrasts with the transcriptional regulation controlling accumulation of CKB protein in postnatal developing cerebellum.

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Tissue- and cell-specific distribution of creatine kinase B: A new and highly specific monoclonal antibody for use in immunohistochemistry

Cited by 57 publications

References 52 publications

Flow of energy in the outer retina in darkness and in light

Flow of energy in the outer retina in darkness and in light

Laser Capture Microdissection-Directed Profiling of Glycolytic and mTOR Pathways in Areas of Selectively Ablated Müller Cells in the Murine Retina

Expression of Creatine Kinase Isoenzyme Genes during Postnatal Development of Rat Brain Cerebrum: Evidence for Posttranscriptional Regulation

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