Center for Integrative Physiology

Edinburgh, United Kingdom

Center for Integrative Physiology

Edinburgh, United Kingdom

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Houston D.A.,Center for Integrative Physiology | Amin A.K.,Royal Infirmary | White T.O.,Royal Infirmary | Smith I.D.M.,Royal Infirmary | Hall A.C.,Center for Integrative Physiology
Osteoarthritis and Cartilage | Year: 2013

Objective: Intra-articular screws are used for internal fixation of osteochondral fragments after fracture or osteochondritis dissecans. This causes cartilage injury potentially leading to chondrocyte death. We have visualised/quantified the hole and zone of cell death (ZCD) in cartilage after drilling/insertion of various articular screws. Method: Using an ex vivo bovine model with transmitted light and confocal laser scanning microscopy (CLSM), the holes and ZCD following drilling/insertion of articular screws (cortical screw, headless variable pitch metallic screw, headless variable pitch bioabsorbable screw) were evaluated. In situ chondrocyte death was determined by live/dead cell viability assay. An imaging/quantification protocol was developed to compare hole diameter and ZCD from drilling/insertion of screws into cartilage. The effect of saline irrigation during drilling on the ZCD was also quantified. Results: Screw insertion created holes in cartilage that were significantly (. P ≤ 0.001) less than the diameters of the equipment used. With a 1.5 mm drill, a ZCD of 580.2 ± 124 μm was produced which increased to 637.0 ± 44 μm following insertion of a 2 mm cortical screw although this was not significant (. P > 0.05). The ZCD from insertion of the variable pitch headless screws (diam. 3.5 mm) was lower for the metallic compared to the bioabsorbable design (800.9 ± 159 vs 1,236.4 ± 212 μm, respectively; P < 0.01). The ZCD from drilling was reduced ~50% (. P < 0.001) by saline irrigation. Conclusions: Cartilage injury during intra-articular screw fixation caused a ZCD around the hole irrespective of screw design. Saline irrigation significantly reduced the ZCD from drilling into cartilage. © 2013 Osteoarthritis Research Society International.


Brose N.,Max Planck Institute for Experimental Medicine | O'Connor V.,University of Southampton | Skehel P.,Center for Integrative Physiology
Biochemical Society Transactions | Year: 2010

Synaptopathy is an increasingly popular term used to define key features of neurodegenerative and psychiatric disease. It implies that disruptions in synaptic structure and function are potentially the major determinant of such brain diseases. The Synaptopathies: Dysfunction of Synaptic Function Biochemical Society Focused Meeting brought together several invited speakers, supplemented with short communications from young scientists, who addressed this possibility. The talks spanned the full gamut of approaches that brought molecular, cellular, systems and whole-animal experimentation together to address how fundamental synaptic biology was increasingly informing on dysfunction in disease. The disease and models thereof discussed included Alzheimer's disease, prions, Huntington's disease, Parkinson's disease, schizophrenia and autism. The audience were asked to reflect on whether synaptopathy, although attractive and conceptually useful, provided a significant explanation as the cause of these major diseases. The breadth of the meeting reinforced the complexity of these brain diseases, supported the significance of synaptic dysfunction in disease, but left open the issue as to whether the prime cause of these disorders could be resolved as simple synaptic dysfunction. Thus, despite revealing a value of synaptopathy, further investigation will be required to reveal its balance in the cause and effect in each of the major brain diseases. ©The Authors.


Ison S.H.,Scottish Agricultural College | D'Eath R.B.,Scottish Agricultural College | Robson S.K.,Scottish Agricultural College | Baxter E.M.,Scottish Agricultural College | And 5 more authors.
Applied Animal Behaviour Science | Year: 2010

In various mammals stress during gestation can result in long term effects on the behaviour and physiology of the offspring. The aims of this study were (i) to characterise individual variation among primiparous sows (gilts) in their behavioural and physiological response to a commercially relevant social challenge (mixing with older sows) during gestation and (ii) to determine whether this variation was associated with a corresponding variation in the subsequent effects on the offspring. After assessing dominance in a food competition test, 24 gilts were studied in groups of six during two one-week periods (unmixed - U). These gilts were split into eight subgroups of three gilts, each of which were mixed with two older sows for one week (mixed - M1) before being returned to their original group of six for two weeks. Mixing was then repeated using different older sows during a second week (mixed - M2). Aggressive behaviour, skin lesions and salivary cortisol increase over baseline were higher, and weight gain was lower during mixed periods. During mixing gilts spent more time in feeder stalls, avoiding sows lying in a straw-bedded area, although there was considerable variation among individuals in the extent to which they interacted with or avoided sows. Gilts which interacted more with sows had lower weight gain and higher lesion scores. Previously dominant gilts had higher salivary cortisol increases during mixing. Measures of the impact of mixing, most notably M1 lesion scores, were associated with the behaviour of their offspring: gilts with more lesions had piglets that were less active and less vocal on the day of weaning and over 31 days post-weaning, and were less aggressive over 31 days post-weaning. Thirty-two piglets (from 16 of the gilts) were either restrained for 30 min or not restrained prior to euthanasia at 60 days of age. The restrained piglets from gilts with higher M1 lesion scores had higher levels of corticotropin-releasing hormone mRNA in the paraventricular nucleus and amygdala. This study demonstrates that differences between individual gilts in their experience of a stressful social challenge co-vary with the extent to which this pre-natal stress impacts on their offspring. © 2010 Elsevier B.V. All rights reserved.


Brunton P.J.,Center for Integrative Physiology | Brunton P.J.,Roslin Institute | Donadio M.V.F.,Center for Integrative Physiology | Donadio M.V.F.,Pontifical Catholic University of Rio Grande do Sul | Russell J.A.,Center for Integrative Physiology
Stress | Year: 2011

We recently reported that male, but not female, offspring born to mothers exposed to social stress during late gestation show heightened anxiety-type behaviour in adulthood. The amygdala organises anxious behaviour, which involves actions of corticotropin-releasing hormone (CRH). CRH gene expression and/or its release are increased in the amygdala in prenatally stressed (PNS) rats. CRH type 1 receptor (CRH-R1) mediates actions of CRH and urocortin I to promote anxiety-like behaviour, whereas the CRH type 2 receptor (CRH-R2) may mediate anxiolytic actions, through actions of urocortins 2 and 3. Here, using quantitative in situ hybridisation, we investigated whether altered CRH receptor mRNA expression in the amygdaloid nuclei may explain the sex differences in anxiety behaviour in adult male and female PNS rats. CRH-R1 mRNA expression was significantly greater in the central amygdala and basolateral amygdala (BLA) in male PNS rats compared with controls, with no change in the basomedial amygdala (BMA) or medial amygdala (MeA). In PNS females, CRH-R1 mRNA expression was greater than controls only in the MeA. Conversely, CRH-R2 mRNA expression was significantly lower in the BMA of male PNS rats compared with controls, but greater in female PNS rats, with no change in the BLA or MeA in either sex. The ratio of CRH-R1:CRH-R2 mRNA in the amygdaloid nuclei was generally increased in PNS males, but not in the PNS females. In conclusion, sex differences in anxiety-type behaviour in PNS rats may be explained by differential mRNA expression for CRH-R1 (pro-anxiogenic) and CRH-R2 (pro-anxiolytic) in the amygdaloid complex. © 2011 Informa Healthcare USA, Inc.


Brydges N.M.,University of Edinburgh | Argyle D.J.,Roslin Institute | Mosley J.R.,Roslin Institute | Duncan J.C.,Roslin Institute | And 3 more authors.
Veterinary Journal | Year: 2012

Dogs with chronic pain have a compromised quality of life. Repeatable and accurate sensory assessments form a means by which the hypersensitivity likely to reflect chronic pain may be quantified. These assessments can be applied to individuals to identify those that may benefit from improved analgesic relief. In this study four sensory assessments were evaluated in dogs presenting with a naturally occurring chronic painful condition (cranial cruciate ligament rupture, CCLR) and were compared with healthy control animals of similar age and weight. Inter-digital von Frey filament and thermal sensitivity tests revealed that the affected hind limb of dogs with CCLR was significantly more sensitive than the opposing limb. Static weight bearing and gait parameter scores were also reduced in the affected hind limb compared to the opposing hind limb of dogs with CCLR; no such differences were found between the hind limbs of healthy (control) dogs. The quantitative sensory tests permitted the differentiation of limbs affected by CCLR from healthy limbs. Dogs presenting with CCLR demonstrate objectively quantitative sensory sensitivities, which may require additional consideration in case management. © 2012 Elsevier Ltd.


Van Breemen C.,University of British Columbia | Fameli N.,University of British Columbia | Evans A.M.,Center for Integrative Physiology
Journal of Physiology | Year: 2013

Abstract This review focuses on how smooth muscle sarcoplasmic reticulum (SR), the major releasable Ca2+ store in these cells, performs its many functions by communicating with the plasma membrane (PM) and other organelles across cytoplasmic nanospaces, defined by membrane-membrane junctions less than 50 nm across. In spite of accumulating evidence in favour of the view that cytoplasmic nanospaces are a prerequisite for effective control of diverse cellular functions, our current understanding of how smooth muscle cells accomplish site- and function-specific Ca2+ signalling remains in its infancy. We first present evidence in support of the view that effective Ca2+ signalling depends on the restricted diffusion of Ca2+ within cytoplasmic nanospaces. We then develop an evidence-based model of the smooth muscle SR - the 'pan-junctional SR' model - that incorporates a network of tubules and quilts that are capable of auto-regulating their Ca2+ content and determining junctional [Ca2+]i through loading and unloading at membrane-membrane nanojunctions. Thereby, we provide a novel working hypothesis in order to inform future investigation into the control of a variety of cellular functions by local Ca2+ signals at junctional nanospaces, from contraction and energy metabolism to nuclear transcription. Based on the current literature, we discuss the molecular mechanisms whereby the SR mediates these multiple functions through the interaction of ion channels and pumps embedded in apposing membranes within inter-organellar junctions. We finally highlight the fact that although most current hypotheses are qualitatively supported by experimental data, solid quantitative simulations are seriously lacking. Considering that at physiological concentrations the number of calcium ions in a typical junctional nanospace between the PM and SR is of the order of 1, ion concentration variability plays a major role as the currency of information transfer and stochastic quantitative modelling will be required to both test and develop working hypotheses. © 2013 The Physiological Society.


Dodson P.D.,Center for Integrative Physiology | Pastoll H.,University of Edinburgh | Nolan M.F.,Center for Integrative Physiology
Journal of Physiology | Year: 2011

The membrane potential dynamics of stellate neurons in layer II of the medial entorhinal cortex are important for neural encoding of location. Previous studies suggest that these neurons generate intrinsic theta-frequency membrane potential oscillations, with a period that depends on neuronal location on the dorsal-ventral axis of the medial entorhinal cortex, and which in behaving animals could support generation of grid-like spatial firing fields. To address the nature and organization of this theta-like activity, we adopt the Lomb method of least-squares spectral analysis. We demonstrate that peaks in frequency spectra that differ significantly from Gaussian noise do not necessarily imply the existence of a periodic oscillator, but can instead arise from filtered stochastic noise or a stochastic random walk. We show that theta-like membrane potential activity recorded from stellate neurons in mature brain slices is consistent with stochastic mechanisms, but not with generation by a periodic oscillator. The dorsal-ventral organization of intrinsic theta-like membrane potential activity, and the modification of this activity during block of HCN channels, both reflect altered frequency distributions of stochastic spectral peaks, rather than tuning of a periodic oscillator. Our results demonstrate the importance of distinguishing periodic oscillations from stochastic processes. We suggest that dorsal-ventral tuning of theta-like membrane potential activity is due to differences in stochastic current fluctuations resulting from organization of ion channels that also control synaptic integration. © 2011 The Authors. Journal compilation © 2011 The Physiological Society.


Greaves J.,Center for Integrative Physiology | Chamberlain L.H.,Center for Integrative Physiology
Journal of Cell Science | Year: 2011

SNAP25 regulates membrane fusion events at the plasma membrane and in the endosomal system, and a functional pool of the protein is delivered to recycling endosomes (REs) and the trans Golgi network (TGN) through an ARF6-dependent cycling pathway. SNAP25 is a peripheral membrane protein, and palmitoylation of a cluster of four cysteine residues mediates its stable association with the membrane. Here, we report that palmitoylation also determines the precise intracellular distribution of SNAP25, and that mutating single palmitoylation sites enhances the amount of SNAP25 at the RE and TGN. The farnesylated CAAX motif from Hras was ligated onto a SNAP25 mutant truncated immediately distal to the cysteine-rich domain. This construct displayed the same intracellular distribution as full-length SNAP25, and decreasing the number of cysteine residues in this construct increased its association with the RE and TGN, confirming the dominant role of the cysteine-rich domain in directing the intracellular distribution of SNAP25. Marked differences in the localisations of SNAP25-CAAX and Hras constructs, each with two palmitoylation sites, were observed, showing that subtle differences in palmitoylated sequences can have a major impact upon intracellular targeting. We propose that the cysteinerich domain of SNAP25 is designed to facilitate the dual function of this SNARE protein at the plasma membrane and endosomes, and that dynamic palmitoylation acts as a mechanism to regulate the precise intracellular patterning of SNAP25. © 2011. Published by The Company of Biologists Ltd.


PubMed | Center for Integrative Physiology
Type: | Journal: European cells & materials | Year: 2016

Partial-thickness cartilage injuries do not heal effectively, potentially leading to degeneration as occurs in post-traumatic osteoarthritis (PTOA). The role of chondrocytes could be crucial in determining the nature of the repair; however, their response to this injury is poorly understood. We have utilised an in vitro bovine osteochondral partial-thickness scalpel injury model and determined chondrocyte properties at and distant from the injury in the presence/absence of (a) serum-free DMEM (340 mOsm), (b) synovial fluid DMEM (SF-DMEM), (c) foetal calf serum DMEM (FCS-DMEM), (d) hyperosmolar serum-free DMEM (600 mOsm), or (e) hyperosmolar FCS-DMEM for up to two weeks. Chondrocytes were fluorescently-labelled with 5-chloromethylfluorescein-diacetate (CMFDA)/propidium iodide (PI) for live/dead cells and imaged using confocal microscopy. Quantitative data were obtained on chondrocyte properties (cell volume, clusters, morphology) at and distant from the injury. In serum-free DMEM, chondrocyte morphology at the injury remained unaffected throughout culture. However, with SF-DMEM or FCS-DMEM the chondrocytes displayed an increase in volume (p < 0.0001), cluster formation (FCS; p < 0.01) and abnormal morphology (p < 0.001) compared to serum-free DMEM. Cluster formation and shape changes during FCS-DMEM culture were more pronounced than with SF-DMEM. SF-DMEM or FCS-DMEM stimulated these changes to chondrocytes at the injury with only small effects on distant cells. Hyperosmolarity inhibited the morphological and volume changes to chondrocytes induced by FCS-DMEM (p < 0.001) and the injured cartilage had the appearance of that in serum-free DMEM. Raised osmolarity may therefore have benefit in preserving the morphological phenotype of chondrocytes at the site of injury, and thus promote more effective integrative repair in partial-thickness cartilage injury.


PubMed | Center for Integrative Physiology
Type: Journal Article | Journal: Osteoarthritis and cartilage | Year: 2013

Intra-articular screws are used for internal fixation of osteochondral fragments after fracture or osteochondritis dissecans. This causes cartilage injury potentially leading to chondrocyte death. We have visualised/quantified the hole and zone of cell death (ZCD) in cartilage after drilling/insertion of various articular screws.Using an exvivo bovine model with transmitted light and confocal laser scanning microscopy (CLSM), the holes and ZCD following drilling/insertion of articular screws (cortical screw, headless variable pitch metallic screw, headless variable pitch bioabsorbable screw) were evaluated. In situ chondrocyte death was determined by live/dead cell viability assay. An imaging/quantification protocol was developed to compare hole diameter and ZCD from drilling/insertion of screws into cartilage. The effect of saline irrigation during drilling on the ZCD was also quantified.Screw insertion created holes in cartilage that were significantly (P0.001) less than the diameters of the equipment used. With a 1.5mm drill, a ZCD of 580.2124m was produced which increased to 637.044m following insertion of a 2mm cortical screw although this was not significant (P>0.05). The ZCD from insertion of the variable pitch headless screws (diam. 3.5mm) was lower for the metallic compared to the bioabsorbable design (800.9159 vs 1,236.4212m, respectively; P<0.01). The ZCD from drilling was reduced 50% (P<0.001) by saline irrigation.Cartilage injury during intra-articular screw fixation caused a ZCD around the hole irrespective of screw design. Saline irrigation significantly reduced the ZCD from drilling into cartilage.

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