Search

Increased levels of lactate, an end-product of glycolysis, have been proposed as a potential surrogate marker for metabolic changes during neuronal excitation. These changes in lactate levels can result in decreased brain pH, which has been implicated in patients with various neuropsychiatric disorders. We previously demonstrated that such alterations are commonly observed in five mouse models of schizophrenia, bipolar disorder, and autism, suggesting a shared endophenotype among these disorders rather than mere artifacts due to medications or agonal state. However, there is still limited research on this phenomenon in animal models, leaving its generality across other disease animal models uncertain. Moreover, the association between changes in brain lactate levels and specific behavioral abnormalities remains unclear. To address these gaps, the International Brain pH Project Consortium investigated brain pH and lactate levels in 109 strains/conditions of 2294 animals with genetic and other experimental manipulations relevant to neuropsychiatric disorders. Systematic analysis revealed that decreased brain pH and increased lactate levels were common features observed in multiple models of depression, epilepsy, Alzheimer’s disease, and some additional schizophrenia models. While certain autism models also exhibited decreased pH and increased lactate levels, others showed the opposite pattern, potentially reflecting subpopulations within the autism spectrum. Furthermore, utilizing large-scale behavioral test battery, a multivariate cross-validated prediction analysis demonstrated that poor working memory performance was predominantly associated with increased brain lactate levels. Importantly, this association was confirmed in an independent cohort of animal models. Collectively, these findings suggest that altered brain pH and lactate levels, which could be attributed to dysregulated excitation/inhibition balance, may serve as transdiagnostic endophenotypes of debilitating neuropsychiatric disorders characterized by cognitive impairment, irrespective of their beneficial or detrimental nature.

Many pathological conditions lead to altered intracellular pH (pHi) disrupting normal cellular functions. The chemical exchange saturation transfer (CEST) method, known as Amine and Amide Concentration Independent Detection (AACID), can produce image contrast that is predominantly dependent on tissue intracellular pHi. The AACID value is linearly related to the ratio of the 3.5 ppm amide CEST effect and the 2.75 ppm amine CEST effect in the physiological range. However, the amine CEST effect at 2 ppm is often more clearly defined in vivo, and may provide greater sensitivity to pH changes. The purpose of the current study was to compare AACID measurement precision utilizing the 2.0 and 2.75 ppm amine CEST effects. We hypothesized that the 2.0 ppm amine CEST resonance would produce measurements with greater sensitivity to pH changes. In the current study, we compare the range of the AACID values obtained in 24 mice with brain tumors and in normal tissue using the 2 ppm and 2.75 ppm amine resonances. All CEST data were acquired on a 9.4T MRI scanner. The AACID measurement range increased by 39% when using the 2 ppm amine resonance compared to the 2.75 ppm resonance, with decreased measurement variability across the brain. These data indicate that in vivo pH measurements made using AACID CEST can be enhanced by incorporating the 2 ppm amine resonance. This approach should be considered for pH measurements made over short intervals when no changes are expected in the concentration of metabolites that contribute to the 2 ppm amine resonance.

Extracellular proton concentration is at 40 nM when pH is 7.4. In disease conditions such as brain ischemia, proton concentration can reach µM range. To respond to this increase in extracellular proton concentration, the mammalian brain expresses at least three classes of proton receptors. Acid-sensing ion channels (ASICs) are the main neuronal cationic proton receptor. The proton-activated chloride channel (PAC), which is also known as (aka) acid-sensitive outwardly rectifying anion channel (ASOR; TMEM206), mediates acid-induced chloride currents. Besides proton-activated channels, GPR4, GPR65 (aka TDAG8, T-cell death-associated gene 8), and GPR68 (aka OGR1, ovarian cancer G protein-coupled receptor 1) function as proton-sensitive G protein-coupled receptors (GPCRs). Though earlier studies on these GPCRs mainly focus on peripheral cells, we and others have recently provided evidence for their functional importance in brain injury. Specifically, GPR4 shows strong expression in brain endothelium, GPR65 is present in a fraction of microglia, while GPR68 exhibits predominant expression in brain neurons. Here, to get a better view of brain acid signaling and its contribution to ischemic injury, we will review the recent findings regarding the differential contribution of proton-sensitive GPCRs to cerebrovascular function, neuroinflammation, and neuronal injury following acidosis and brain ischemia. [ABSTRACT FROM AUTHOR]

Many pathological conditions lead to altered intracellular pH (pHi) disrupting normal cellular functions. The chemical exchange saturation transfer (CEST) method, known as Amine and Amide Concentration Independent Detection (AACID), can produce image contrast that is predominantly dependent on tissue intracellular pHi. The AACID value is linearly related to the ratio of the 3.5 ppm amide CEST effect and the 2.75 ppm amine CEST effect in the physiological range. However, the amine CEST effect at 2 ppm is often more clearly defined in vivo, and may provide greater sensitivity to pH changes. The purpose of the current study was to compare AACID measurement precision utilizing the 2.0 and 2.75 ppm amine CEST effects. We hypothesized that the 2.0 ppm amine CEST resonance would produce measurements with greater sensitivity to pH changes. In the current study, we compare the range of the AACID values obtained in 24 mice with brain tumors and in normal tissue using the 2 ppm and 2.75 ppm amine resonances. All CEST data were acquired on a 9.4T MRI scanner. The AACID measurement range increased by 39% when using the 2 ppm amine resonance compared to the 2.75 ppm resonance, with decreased measurement variability across the brain. These data indicate that in vivo pH measurements made using AACID CEST can be enhanced by incorporating the 2 ppm amine resonance. This approach should be considered for pH measurements made over short intervals when no changes are expected in the concentration of metabolites that contribute to the 2 ppm amine resonance. [ABSTRACT FROM AUTHOR]

Increased levels of lactate, an end-product of glycolysis, have been proposed as a potential surrogate marker for metabolic changes during neuronal excitation. These changes in lactate levels can result in decreased brain pH, which has been implicated in patients with various neuropsychiatric disorders. We previously demonstrated that such alterations are commonly observed in five mouse models of schizophrenia, bipolar disorder, and autism, suggesting a shared endophenotype among these disorders rather than mere artifacts due to medications or agonal state. However, there is still limited research on this phenomenon in animal models, leaving its generality across other disease animal models uncertain. Moreover, the association between changes in brain lactate levels and specific behavioral abnormalities remains unclear. To address these gaps, the International Brain pH Project Consortium investigated brain pH and lactate levels in 109 strains/conditions of 2294 animals with genetic and other experimental manipulations relevant to neuropsychiatric disorders. Systematic analysis revealed that decreased brain pH and increased lactate levels were common features observed in multiple models of depression, epilepsy, Alzheimer's disease, and some additional schizophrenia models. While certain autism models also exhibited decreased pH and increased lactate levels, others showed the opposite pattern, potentially reflecting subpopulations within the autism spectrum. Furthermore, utilizing large-scale behavioral test battery, a multivariate cross-validated prediction analysis demonstrated that poor working memory performance was predominantly associated with increased brain lactate levels. Importantly, this association was confirmed in an independent cohort of animal models. Collectively, these findings suggest that altered brain pH and lactate levels, which could be attributed to dysregulated excitation/inhibition balance, may serve as transdiagnostic endophenotypes of debilitating neuropsychiatric disorders characterized by cognitive impairment, irrespective of their beneficial or detrimental nature., Competing Interests: HH, HS, SH, GS, YT, MT, MI, MS, IH, KH, MH, AN, YM, SO, MM, AU, YK, AM, KN, SK, TS, TI, HN, YG, MR, TT, KY, NT, HK, JI, IN, TK, TT, MD, HN, KT, KS, KN, TT, HF, YS, KY, LR, NH, HM, TY, SW, TA, KT, TN, HK, FH, AH, HH, KT, TT, TK, TK, IG, GC, NA, HH, SK, TK, MH, YH, RM, KO, AH, LB, IY, YH, HK, YM, KI, HK, BK, HO, YY, AT, NK, SG, MI, MN, KM, EO, IO, TS, KY, HA, KT, KI, TS, AK, MF, NU, TK, TT, YY, MI, TH, KI, KN, SH, KE, KH, YT, SN, NS, TY, NU, TY, YK, RO, NS, KS, SH, KO, HM, TF, MT, HS, DS, YI, TI, YK, TY, KS, TM No competing interests declared, SY, NN Employee of Takeda Pharmaceutical Company, Ltd

Background: PINK1 mutations are likely to affect mitochondrial function. The objective of this study was to study brain mitochondrial function in patients with early-onset Parkinson's disease, with or without PINK1 mutations.Methods: We investigated brain intracellular pH, mitochondrial activity, and energetics with functional magnetic resonance spectroscopy in patients with early-onset Parkinson's disease with PINK1 mutations (n = 10), early-onset Parkinson's disease without PINK1 mutations (n = 10), and healthy sex- and age-matched subjects (n = 20). We measured peak areas of phosphocreatine and beta adenosine triphosphate.Results: The EOPD- group had normal PCr + βATP contents at rest (P = NS) and under activation (P = NS), but reduced contents during recovery (P < 0.001). The EOPD+ group had abnormal PCr + βATP contents at rest (P < 0.001) and during activation (P < 0.001); during recovery, the contents only partially recovered (P < 0.001). Brain intracellular pH alterations were more severe with EOPD+ than with EOPD-.Conclusions: Brain mitochondrial impairments were similar in early-onset Parkinson's disease without PINK1 mutations and late-onset Parkinson's disease. However, mitochondrial impairments were more severe in early-onset Parkinson's disease with PINK1 mutations. © 2020 International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]

Microglial associations with both the major Alzheimer's disease (AD) pathognomonic entities, β‐amyloid‐positive plaques and tau‐positive neurofibrillary tangles, have been noted in previous investigations of both human tissue and mouse models. However, the precise nature of their role in the pathogenesis of AD is debated; the major working hypothesis is that pro‐inflammatory activities of activated microglia contribute to disease progression. In contrast, others have proposed that microglial dystrophy with a loss of physiological and neuroprotective activities promotes neurodegeneration. This immunohistochemical study sought to gain clarity in this area by quantifying the morphological subtypes of microglia in the mildly‐affected primary visual cortex (PVC), the moderately affected superior frontal cortex (SFC) and the severely affected inferior temporal cortex (ITC) of 8 AD cases and 15 age and gender‐matched, non‐demented controls with ranging AD‐type pathology. AD cases had increased β‐amyloid and tau levels compared to controls in all regions. Neuronal loss was observed in the SFC and ITC, and was associated with atrophy in the latter. A major feature of the ITC in AD was a decrease in ramified (healthy) microglia with image analysis confirming reductions in arborized area and skeletal complexity. Activated microglia were not associated with AD but were increased in non‐demented controls with greater AD‐type pathology. Microglial clusters were occasionally associated with β‐amyloid‐ and tau‐positive plaques but represented less than 2% of the total microglial population. Dystrophic microglia were not associated with AD, but were inversely correlated with brain pH suggesting that agonal events were responsible for this morphological subtype. Overall these novel findings suggest that there is an early microglial reaction to AD‐type pathology but a loss of healthy microglia is the prominent feature in severely affected regions of the AD brain. [ABSTRACT FROM AUTHOR]

Postmortem studies reveal that the brain pH in schizophrenia patients is lower than normal. The exact cause of this low pH is unclear, but increased lactate levels due to abnormal energy metabolism appear to be involved. Schizophrenia patients display distinct changes in mitochondria number, morphology, and function, and such changes promote anaerobic glycolysis, elevating lactate levels. pH can affect neuronal activity as H+ binds to numerous proteins in the nervous system and alters the structure and function of the bound proteins. There is growing evidence of pH change associated with cognition, emotion, and psychotic behaviors. Brain has delicate pH regulatory mechanisms to maintain normal pH in neurons/glia and extracellular fluid, and a change in these mechanisms can affect, or be affected by, neuronal activities associated with schizophrenia. In this review, we discuss the current understanding of the cause and effect of decreased brain pH in schizophrenia based on postmortem human brains, animal models, and cellular studies. The topic includes the factors causing decreased brain pH in schizophrenia, mitochondria dysfunction leading to altered energy metabolism, and pH effects on the pathophysiology of schizophrenia. We also review the acid/base transporters regulating pH in the nervous system and discuss the potential contribution of the major transporters, sodium hydrogen exchangers (NHEs), and sodium-coupled bicarbonate transporters (NCBTs), to schizophrenia.

The choroid plexus epithelium (CPE) is located in the ventricular system of the brain, where it secretes the majority of the cerebrospinal fluid (CSF) that fills the ventricular system and surrounds the central nervous system. The CPE is a highly vascularized single layer of cuboidal cells with an unsurpassed transepithelial water and solute transport rate. Several members of the slc4a family of bicarbonate transporters are expressed in the CPE. In the basolateral membrane the electroneutral Na+ dependent Cl-/HCO3- exchanger, NCBE (slc4a10) is expressed. In the luminal membrane, the electrogenic Na+:HCO3- cotransporter, NBCe2 (slc4a5) is expressed. The electroneutral Na+:HCO3- cotransporter, NBCn1 (slc4a7), has been located in both membranes. In addition to the bicarbonate transporters, the Na+/H+ exchanger, NHE1 (slc9a1), is located in the luminal membrane of the CPE. Genetically modified mice targeting slc4a2, slc4a5, Slc4a7, Slc4a10, and Slc9a1 have been generated. Deletion of slc4a5, 7 or 10, or slc9a1 has numerous impacts on CP function and structure in these mice. Removal of the transporters affects brain ventricle size (slc4a5 and slc4a10) and intracellular pH regulation (slc4a7 and slc4a10). In some instances, removal of the proteins from the CPE (slc4a5, 7 and 10) causes changes in abundance and localization of non-target transporters known to be involved in pH regulation and CSF secretion. The focus of this review is to combine the insights gathered from these knockout mice to highlight the impact of slc4 gene deletion on the CSF production and intracellular pH regulation resulting from the deletion of slc4a5, 7 and 10, and slc9a1. Furthermore, the review contains a comparison of the described human mutations of these genes to the findings in the knockout studies. Finally, the future perspective of utilizing these proteins as potential targets for the treatment of CSF disorders will be discussed.

Panic attacks (PAs), the core feature of panic disorder, represent a common phenomenon in the general adult population and are associated with a considerable decrease in quality of life and high health care costs. To date, the underlying pathophysiology of PAs is not well understood. A unique feature of PAs is that they represent a rare example of a psychopathological phenomenon that can be reliably modeled in the laboratory in panic disorder patients and healthy volunteers. The most effective techniques to experimentally trigger PAs are those that acutely disturb the acid–base homeostasis in the brain: inhalation of carbon dioxide (CO 2 ), hyperventilation, and lactate infusion. This review particularly focuses on the use of CO 2 inhalation in humans and rodents as an experimental model of panic. Besides highlighting the different methodological approaches, the cardio-respiratory and the endocrine responses to CO 2 inhalation are summarized. In addition, the relationships between CO 2 level, changes in brain pH, the serotonergic system, and adaptive physiological and behavioral responses to CO 2 exposure are presented. We aim to present an integrated psychological and neurobiological perspective. Remaining gaps in the literature and future perspectives are discussed. [ABSTRACT FROM AUTHOR]

Abstract: Recent experiments suggest that T1 relaxation in the rotating frame (T1ρ) detects localized metabolic changes in the human visual cortex induced by a flashing checkerboard task. Possible sources of the T1ρ signal include pH, glucose, and glutamate concentrations as well as changes in cerebral blood volume. In this study we explored the relationship of the T1ρ signal changes related to cerebral blood volume changes by employing inferior saturation pulses. Our hypothesis was that there would be a contribution of cerebral blood volume to the functional T1ρ signal, but a majority of the signal would correspond to metabolic changes. In addition, the relationship between T1ρ and pH was explored by manipulating the frequency of the flashing checkerboard and imaging with T1ρ, BOLD, and 31P spectroscopy. We hypothesized that T1ρ and pH changes would be sensitive to the stimulation frequency. To test this hypothesis, we used a full-field visual flashing checkerboard and varied the frequency between 1, 4, and 7Hz. Supporting our hypotheses, we found that approximately 73% of the measured signal change corresponds to metabolism in vivo and that increasing stimulation frequency increased responses measured by all three imaging modalities. The activation area detected by T1ρ overlapped to a large degree with that detected by BOLD, although the T1ρ response area was significantly smaller. 31P spectroscopy detected a greater acidosis with the higher stimulation frequencies. These observations suggest that, similar to the BOLD response, the magnitude of the T1ρ and pH response depends on stimulation frequency and is thus likely to be activity-dependent. [Copyright &y& Elsevier]

Background: Abnormal brain pH has been suggested to play a critical role in panic disorder. To investigate this possibility, we employed a pH-sensitive magnetic resonance (MR) imaging strategy (T1 relaxation in the rotating frame [T1ρ]) and conventional blood oxygen level-dependent (BOLD) imaging. Methods: Thirteen panic disorder participants and 13 matched control subjects were enrolled in the study. T1ρ and BOLD were used to study the functional response to a visual flashing checkerboard and their relationship to panic symptoms assessed using the Beck Anxiety Inventory. Results: In response to visual stimulation, T1ρ imaging revealed a significantly greater increase in the visual cortex of panic disorder participants. T1ρ also detected a stimulus-evoked decrease in the anterior cingulate cortex. Blood oxygen level-dependent imaging detected no functional differences between groups. The correspondence between panic symptoms and functional T1ρ response identified significant relationships within the left inferior parietal lobe, left middle temporal gyrus, and right insula. No relationships were found between panic symptoms and the BOLD signal. Conclusions: The data suggest greater activity-evoked T1ρ changes in the visual cortex and anterior cingulate cortex of panic disorder participants. These observations are consistent with a pH dysregulation in panic disorder. In addition, our data suggest that T1ρ imaging may provide information about panic disorder that is distinct from conventional BOLD imaging and may reflect abnormalities in pH and/or brain metabolism. [ABSTRACT FROM AUTHOR]

Purpose: To use (13) C magnetic resonance spectroscopy (MRS) and (31) P MRS to develop a direct assay for regional [HCO3-] in the human brain and to define brain pH and physiological response of [HCO3-] to fasting.Materials and Methods: Seven healthy subjects underwent MRS examinations on a 1.5T MRI scanner. Subjects were well fed with repeated examinations performed after 4 and 12 hours of fasting. Proton noise decoupling (13) C MRS were acquired using pulse and acquired acquisition while (31) P MRS were acquired using a 2D chemical shift imaging method with relaxation time (TR) of 2 seconds.Results: Fasting brain bicarbonate concentrations (6.7 ± 2.5 mM for 12-hour fasting, P = 0.002 and 8.3 ± 2.1 mM for 4-hour fasting, P = 0.015) are significantly reduced compared to fed state (11.6 ± 1.3 mM). However, no significant difference in brain pH was observed, confirming the critical role of pCO2 in intracerebral pH homeostasis.Conclusion: We demonstrated that the intracellular HCO3- in human brain is readily modified by diet but appears to have no measurable effect on cerebral pH. Natural abundance (13) C can provide useful information relevant to human brain pH homeostasis by providing information for HCO3-. [ABSTRACT FROM AUTHOR]

Purpose To use 13C magnetic resonance spectroscopy (MRS) and 31P MRS to develop a direct assay for regional [HCO3−] in the human brain and to define brain pH and physiological response of [HCO3−] to fasting. Materials and Methods Seven healthy subjects underwent MRS examinations on a 1.5T MRI scanner. Subjects were well fed with repeated examinations performed after 4 and 12 hours of fasting. Proton noise decoupling 13C MRS were acquired using pulse and acquired acquisition while 31P MRS were acquired using a 2D chemical shift imaging method with relaxation time (TR) of 2 seconds. Results Fasting brain bicarbonate concentrations (6.7 ± 2.5 mM for 12-hour fasting, P = 0.002 and 8.3 ± 2.1 mM for 4-hour fasting, P = 0.015) are significantly reduced compared to fed state (11.6 ± 1.3 mM). However, no significant difference in brain pH was observed, confirming the critical role of pCO2 in intracerebral pH homeostasis. Conclusion We demonstrated that the intracellular HCO3− in human brain is readily modified by diet but appears to have no measurable effect on cerebral pH. Natural abundance 13C can provide useful information relevant to human brain pH homeostasis by providing information for HCO3−. J. Magn. Reson. Imaging 2014;39:398-401. © 2013 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Encephalopathy consequent on perinatal hypoxia-ischemia occurs in 1-3 per 1,000 term births in the UK and frequently leads to serious and tragic consequences that devastate lives and families, with huge financial burdens for society. Although the recent introduction of cooling represents a significant advance, only 40 % survive with normal neurodevelopmental function. There is thus a significant unmet need for novel, safe, and effective therapies to optimize brain protection following brain injury around birth. The Na/H exchanger (NHE) is a membrane protein present in many mammalian cell types. It is involved in regulating intracellular pH and cell volume. NHE1 is the most abundant isoform in the central nervous system and plays a role in cerebral damage after hypoxia-ischemia. Excessive NHE activation during hypoxia-ischemia leads to intracellular Na overload, which subsequently promotes Ca entry via reversal of the Na/Ca exchanger. Increased cytosolic Ca then triggers the neurotoxic cascade. Activation of NHE also leads to rapid normalization of pH and an alkaline shift in pH. This rapid recovery of brain intracellular pH has been termed pH paradox as, rather than causing cells to recover, this rapid return to normal and overshoot to alkaline values is deleterious to cell survival. Brain pH changes are closely involved in the control of cell death after injury: an alkalosis enhances excitability while a mild acidosis has the opposite effect. We have observed a brain alkalosis in 78 babies with neonatal encephalopathy serially studied using phosphorus-31 magnetic resonance spectroscopy during the first year after birth (151 studies throughout the year including 56 studies of 50 infants during the first 2 weeks after birth). An alkaline brain pH was associated with severely impaired outcome; the degree of brain alkalosis was related to the severity of brain injury on MRI and brain lactate concentration; and a persistence of an alkaline brain pH was associated with cerebral atrophy on MRI. Experimental animal models of hypoxia-ischemia show that NHE inhibitors are neuroprotective. Here, we review the published data on brain pH in neonatal encephalopathy and the experimental studies of NHE inhibition and neuroprotection following hypoxia-ischemia. [ABSTRACT FROM AUTHOR]

The choroid plexus epithelium (CPE) is located in the ventricular system of the brain, where it secretes the majority of the cerebrospinal fluid (CSF) that fills the ventricular system and surrounds the central nervous system. The CPE is a highly vascularized single layer of cuboidal cells with an unsurpassed transepithelial water and solute transport rate. Several members of the slc4a family of bicarbonate transporters are expressed in the CPE. In the basolateral membrane the electroneutral Na+ dependent Cl-/HCO3 - exchanger, NCBE (slc4a10) is expressed. In the luminal membrane, the electrogenic Na+:HCO3 - cotransporter, NBCe2 (slc4a5) is expressed. The electroneutral Na+:HCO3 - cotransporter, NBCn1 (slc4a7), has been located in both membranes. In addition to the bicarbonate transporters, the Na+/H+ exchanger, NHE1 (slc9a1), is located in the luminal membrane of the CPE. Genetically modified mice targeting slc4a2, slc4a5, slc4a7, slc4a10, and slc9a1 have been generated. Deletion of slc4a5, 7 or 10, or slc9a1 has numerous impacts on CP function and structure in these mice. Removal of the transporters affects brain ventricle size (slc4a5 and slc4a10) and intracellular pH regulation (slc4a7 and slc4a10). In some instances, removal of the proteins from the CPE (slc4a5, 7, and 10) causes changes in abundance and localization of non-target transporters known to be involved in pH regulation and CSF secretion. The focus of this review is to combine the insights gathered from these knockout mice to highlight the impact of slc4 gene deletion on the CSF production and intracellular pH regulation resulting from the deletion of slc4a5, 7 and 10, and slc9a1. Furthermore, the review contains a comparison of the described human mutations of these genes to the findings in the knockout studies. Finally, the future perspective of utilizing these proteins as potential targets for the treatment of CSF disorders will be discussed. [ABSTRACT FROM AUTHOR]

Tissue quality control measures are routinely performed in brain banks with the assessment of brain pH being the most common measure. In some brain banks the assessment of the RNA integrity number is also performed, although this requires access to specialised equipment and is more expensive. The aim of this study is to determine if there is a correlation between the visual assessment of cerebellar granule cell integrity and brain pH or RIN. One hundred and five consecutive cases from the NSW Tissue Resource Centre, Sydney, Australia were accessed. The cerebrum was hemisected and one hemisphere sliced parasagittally at approximately 1-2 cm intervals and frozen. The other hemisphere was fixed in 15% buffered formalin for 2-3 weeks. The contralateral cerebellar hemisphere was preserved in the same manner as the cerebral hemisphere. Samples of fixed tissue were embedded in paraffin, 7 μm sections cut and stained routinely with hematoxylin and eosin. The granular cell layer (GCL) was assessed microscopically to determine the degree of autolytic degradation. Degradation was graded as nil, mild, moderate or severe. Brain tissue pH and RIN were measured using standardised protocols. This study showed that both brain pH and RIN significantly correlated with the severity of the degradation of the cerebellar granule cell layer. This additional screening tool can be performed during routine histological review of the cerebellar tissue to assess the suitability for further investigation of tissue quality. [ABSTRACT FROM AUTHOR]

Localized pH changes have been suggested to occur in the brain during normal function. However, the existence of such pH changes has also been questioned. Lack of methods for noninvasively measuring pH with high spatial and temporal resolution has limited insight into this issue. Here we report that a magnetic resonance imaging (MRI) strategy, T1 relaxation in the rotating frame (T1ρ), is sufficiently sensitive to detect widespread pH changes in the mouse and human brain evoked by systemically manipulating carbon dioxide or bicarbonate. Moreover, T1ρ detected a localized acidosis in the human visual cortex induced by a flashing checkerboard. Lactate measurements and pH-sensitive 31P spectroscopy at the same site also identified a localized acidosis. Consistent with the established role for pH in blood flow recruitment, T1? correlated with blood oxygenation level-dependent contrast commonly used in functional MRI. However, T1? was not directly sensitive to blood oxygen content. These observations indicate that localized pH fluctuations occur in the human brain during normal function. Furthermore, they suggest a unique functional imaging strategy based on pH that is independent of traditional functional MRI contrast mechanisms. [ABSTRACT FROM AUTHOR]

Abstract: The quality of results from microarray studies depends on RNA quality, which can be significantly influenced by postmortem factors. The aim of this study was to determine which postmortem factors and/or RNA electropherogram characteristics best correspond to microarray output and can be used to prospectively screen RNA prior to microarray analysis. Total RNA was extracted (N =125) from gray and white matter of postmortem frontal and occipital lobe tissue, acquired from normal controls, and patients with schizophrenia, bipolar disorder or major depression. Electropherograms were generated by the Agilent BioAnalyzer 2100, allowing calculation of the 28S/18S ratio, the 18S/baseline peak ratio and the RNA Integrity Number (RIN). These values were compared to post-hybridization image analysis of Affymetrix microarrays. The postmortem variables correlated with some quality measures but could not be used as effective screening tools. Logistic regression demonstrated that all three electropherogram measures were predictive for microarray quality, and that the RIN threshold predictive of “good quality” (>35% present calls) was most consistent with that of prior studies. The optimal RIN must be determined by the investigator''s specifications for false inclusion and false exclusion. In contrast to RIN, the quality threshold for the 28S/18S ratio has proven unacceptably variable, due to sensitivity to slight differences in protocol and/or tissue source. In conclusion, the measures we found useful as screening criteria do not replace the need to exclude samples after a microarray analysis is performed, as an acceptable percent call rate and other measures of microarray quality represent the desired endpoint. [Copyright &y& Elsevier]

Abstract: Aims: To determine whether there are factors during apparent rapid heroin overdose death that affect agonal state and thus brain pH (index of hypoxia) that can influence neurobiological systems linked to drug abuse. Design and methods: Brain specimens and autopsy/medical reports were investigated in subjects dying from heroin overdose (n =70) and compared to normal controls (n =45) as well as suicide victims (n =31) with a documented rapid cause of death. Detailed autopsy material was characterized as to positive and negative respiratory distress in relation to brain pH; drug toxicity and other demographic information was also evaluated. In situ hybridization histochemistry was used to study mRNA expression levels of dopamine (e.g., D2 receptor, dopamine transporter) and opioid (e.g., proenkephalin) related markers in various structures in relation to brain pH. Findings: Brain pH was generally reduced in heroin overdose cases versus normal and suicide subjects. There was, however, significant variation in heroin overdose deaths related to differences in respiratory distress that differentially altered brain pH levels. Various factors such as vomit inhalation, resuscitation, pulmonary embolism and suffocation contributed to positive respiratory distress. Elevated brain pH was observed in heroin overdose with positive alcohol toxicity suggesting potentiated alcohol-induced rapidity of heroin deaths. mRNA expression levels of the dopamine-related genes and proenkephalin were positively correlated with brain pH. Conclusions: Respiratory distress contributes to variations in the acute agonal state during heroin overdose death that differentially alters brain pH levels and significantly impacts mRNA levels. Such findings should be considered for postmortem molecular/neurochemical neurobiological studies of opiate abusers. [Copyright &y& Elsevier]

Many pathological conditions lead to altered intracellular pH (pH i ) disrupting normal cellular functions. The chemical exchange saturation transfer (CEST) method, known as Amine and Amide Concentration Independent Detection (AACID), can produce image contrast that is predominantly dependent on tissue intracellular pH i . The AACID value is linearly related to the ratio of the 3.5 ppm amide CEST effect and the 2.75 ppm amine CEST effect in the physiological range. However, the amine CEST effect at 2 ppm is often more clearly defined in vivo, and may provide greater sensitivity to pH changes. The purpose of the current study was to compare AACID measurement precision utilizing the 2.0 and 2.75 ppm amine CEST effects. We hypothesized that the 2.0 ppm amine CEST resonance would produce measurements with greater sensitivity to pH changes . In the current study, we compare the range of the AACID values obtained in 24 mice with brain tumors and in normal tissue using the 2 ppm and 2.75 ppm amine resonances. All CEST data were acquired on a 9.4T MRI scanner. The AACID measurement range increased by 39% when using the 2 ppm amine resonance compared to the 2.75 ppm resonance, with decreased measurement variability across the brain. These data indicate that in vivo pH measurements made using AACID CEST can be enhanced by incorporating the 2 ppm amine resonance. This approach should be considered for pH measurements made over short intervals when no changes are expected in the concentration of metabolites that contribute to the 2 ppm amine resonance.

Postmortem studies reveal that the brain pH in schizophrenia patients is lower than normal. The exact cause of this low pH is unclear, but increased lactate levels due to abnormal energy metabolism appear to be involved. Schizophrenia patients display distinct changes in mitochondria number, morphology, and function, and such changes promote anaerobic glycolysis, elevating lactate levels. pH can affect neuronal activity as H+ binds to numerous proteins in the nervous system and alters the structure and function of the bound proteins. There is growing evidence of pH change associated with cognition, emotion, and psychotic behaviors. Brain has delicate pH regulatory mechanisms to maintain normal pH in neurons/glia and extracellular fluid, and a change in these mechanisms can affect, or be affected by, neuronal activities associated with schizophrenia. In this review, we discuss the current understanding of the cause and effect of decreased brain pH in schizophrenia based on postmortem human brains, animal models, and cellular studies. The topic includes the factors causing decreased brain pH in schizophrenia, mitochondria dysfunction leading to altered energy metabolism, and pH effects on the pathophysiology of schizophrenia. We also review the acid/base transporters regulating pH in the nervous system and discuss the potential contribution of the major transporters, sodium hydrogen exchangers (NHEs), and sodium-coupled bicarbonate transporters (NCBTs), to schizophrenia. [ABSTRACT FROM AUTHOR]

• Brain pH was determined with CEST MRI in lithium-pilocarpine rats with seizures. • pH was alkaline in the hippocampus and piriform cortex compared to controls. • The alkaline shift was marginal in the thalamus. • The alkaline shift was not significant in the amygdala and cerebral cortex. • Brain pH in patients with chronic seizures might be similarly alkaline shifted. Brain pH is thought to be important in epilepsy. The regulation of brain pH is, however, still poorly understood in animal models of chronic seizures (SZ) as well as in patients with intractable epilepsy. We used chemical exchange saturation transfer (CEST) MRI to noninvasively determine if the pH is alkaline shifted in a rodent model of the mesial temporal lobe (MTL) epilepsy with chronic SZ. Taking advantage of its high spatial resolution, we determined the pH values in specific brain regions believed to be important in this model produced by lithium-pilocarpine injection. All animals developed status epilepticus within 90 min after the lithium-pilocarpine administration, but one animal died within 24 hrs. All the surviving animals developed chronic SZ during the first 2 months. After SZ developed, brain pH was determined in the pilocarpine and control groups (n = 8 each). Epileptiform activity was documented in six pilocarpine rats with scalp EEG. The brain pH was estimated using two methods based on magnetization transfer asymmetry and amide proton transfer ratio. The pH was alkaline shifted in the pilocarpine rats (one outlier excluded) compared to the controls in the hippocampus (7.29 vs 7.17, t -test, p < 0.03) and the piriform cortex (7.34 vs. 7.06, p < 0.005), marginally more alkaline in the thalamus (7.13 vs. 7.01, p < 0.05), but not in the cerebral cortex (7.18 vs. 7.08, p > 0.05). Normalizing the brain pH may lead to an effective non-surgical method for treating intractable epilepsy as it is known that SZ can be eliminated by lowering the pH. [ABSTRACT FROM AUTHOR]

The fraction of the cardiac output distributed to the brain was studied with a particle distribution method at different acutely induced arterial P levels in control rats and in rats kept during 24 h before the experiment in an atmosphere containing either 10% CO in air (prolonged hypercapnia) or 10% O in N (hypoxic hyperventilation producing prolonged hypocapnia). The curve relating the fraction of the cardiac output distributed to the brain and the P is shifted upwards and to the left after prolonged hypocapnia, and downwards and to the right after prolonged hypercapnia. A similar shift is observed when orthosympathetic vasoconstrictor activity is eliminated in the three groups of rats by alpha-adrenergic receptor blockade. The changes in the fraction of the cardiac output perfusing the brain are essentially due to modifications in the cerebral blood flow, since the cardiac output was not significantly influenced by the alterations in the P. The results are discussed in relation to the changes occurring in the CO/HCO dissociation curve in blood and brain during prolonged respiratory changes in the acid-base balance. [ABSTRACT FROM AUTHOR]

A detailed description is given of a new model of reversible focal cerebral ischaemia in rats. A spherical embolus, attached to the end of an 8-0 surgical thread was injected into the middle cerebral artery (MCA) via the external and internal carotid arteries (ECA and ICA) and could be withdrawn by pulling the extravascular portion of the thread when reperfusion was to follow ischaemia. In contrast to similar techniques, the 8-0 thread did not block blood flow in either the common carotid artery (CCA) or the ICA during ischaemia and, as the CCA did not have to be ligated, flow could be restored via the ipsilateral CCA and ICA after the ischaemic period. Neurological deficit, mortality rate, tissue water content, regional pH, ATP and, in some experiments, electroencephalogram (EEG) and cerebral blood flow (CBF) were evaluated in ischaemia and after various length of reperfusion. The overall mortality rate was 21%; all these animals were lost in the first hours of recirculation. The water content of their brains differed significantly from those who survived 24 hours (81 and 77% respectively). A significant difference in CBF of the ipsi- and contralateral putamen was found immediately after embolization (a CBF decrease of 26±5 and 5±5 ml/100 g/min, respectively), and relative hyperperfusion (+23 ±27 ml/100 g/min) was observed in the ipsilateral putamen during reperfusion. EEG amplitude declined on both the affected and non-affected sides after embolization (to 54±8% and 71 ±6% of pre-ischaemic values) and remained decreased (66 ±8% of control) on the ipsilateral side after 1 hour of reperfusion. Embolization resulted in tissue acidosis and ATP depletion predominantly in the somatosensory cortex, caudoto-putamen, internal capsule, anterior thalamus and hippocampus. The spatial extent of metabolic alterations increased in the first two hours of recirculation, then decreased by 24 hours, together with the improvement of clinical signs. Providing a better model for human embolic strokes this model is suggested for studying consequences of reversible focal ischaemia. [ABSTRACT FROM AUTHOR]

African naked mole-rats were likely the first mammals to evolve eusociality, and thus required adaptations to conserve energy and tolerate the low oxygen (O 2) and high carbon dioxide (CO 2) of a densely populated fossorial nest. As hypercapnia is known to suppress neuronal activity, we studied whether naked mole-rats might demonstrate energy savings in GABAergic inhibition. Using whole-colony behavioral monitoring of captive naked mole-rats, we found a durable nest, characterized by high CO 2 levels, where all colony members spent the majority of their time. Analysis of the naked mole-rat genome revealed, uniquely among mammals, a histidine point variation in the neuronal potassium-chloride cotransporter 2 (KCC2). A histidine missense substitution mutation at this locus in the human ortholog of KCC2, found previously in patients with febrile seizures and epilepsy, has been demonstrated to diminish neuronal Cl− extrusion capacity, and thus impairs GABAergic inhibition. Seizures were observed, without pharmacological intervention, in adult naked mole-rats exposed to a simulated hyperthermic surface environment, causing systemic hypocapnic alkalosis. Consistent with the diminished function of KCC2, adult naked mole-rats demonstrate a reduced efficacy of inhibition that manifests as triggering of seizures at room temperature by the GABA A receptor (GABA A R) positive allosteric modulator diazepam. These seizures are blocked in the presence of nest-like levels of CO 2 and likely to be mediated through GABA A R activity, based on in vitro recordings. Thus, altered GABAergic inhibition adds to a growing list of adaptations in the naked mole-rat and provides a plausible proximate mechanism for nesting behavior, where a return to the colony nest restores GABA-mediated inhibition. • Eusocial African naked mole-rats raise colony-nest CO 2 to anticonvulsant levels • Naked mole-rats harbor an epilepsy-related variant in neuronal Cl– extruder KCC2 • GABA-acting antiseizure drug diazepam triggers seizures in naked mole-rats • Diazepam is rendered seizure-suppressing in nest-like levels of CO 2 in inhaled air Zions et al. show a dependence of captive naked mole-rats on elevated carbon dioxide (CO 2) in the colony nest to avoid seizure. They also show a species-specific mutation of the cation-chloride cotransporter KCC2, compromising neuronal Cl– extrusion. Evolution may have favored selection of this energy-saving mutation, which is masked by the anticonvulsant properties of CO 2. [ABSTRACT FROM AUTHOR]

Recent experiments suggest that T1 relaxation in the rotating frame (T1ρ) detects localized metabolic changes in the human visual cortex induced by a flashing checkerboard task. Possible sources of the T1ρ signal include pH, glucose, and glutamate concentrations as well as changes in cerebral blood volume. In this study we explored the relationship of the T1ρ signal changes related to cerebral blood volume changes by employing inferior saturation pulses. Our hypothesis was that there would be a contribution of cerebral blood volume to the functional T1ρ signal, but a majority of the signal would correspond to metabolic changes. In addition, the relationship between T1ρ and pH was explored by manipulating the frequency of the flashing checkerboard and imaging with T1ρ, BOLD, and (31)P spectroscopy. We hypothesized that T1ρ and pH changes would be sensitive to the stimulation frequency. To test this hypothesis, we used a full-field visual flashing checkerboard and varied the frequency between 1, 4, and 7Hz. Supporting our hypotheses, we found that approximately 73% of the measured signal change corresponds to metabolism in vivo and that increasing stimulation frequency increased responses measured by all three imaging modalities. The activation area detected by T1ρ overlapped to a large degree with that detected by BOLD, although the T1ρ response area was significantly smaller. (31)P spectroscopy detected a greater acidosis with the higher stimulation frequencies. These observations suggest that, similar to the BOLD response, the magnitude of the T1ρ and pH response depends on stimulation frequency and is thus likely to be activity-dependent., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The choroid plexus epithelium (CPE) is located in the ventricular system of the brain, where it secretes the majority of the cerebrospinal fluid (CSF) that fills the ventricular system and surrounds the central nervous system. The CPE is a highly vascularized single layer of cuboidal cells with an unsurpassed transepithelial water and solute transport rate. Several members of the slc4a family of bicarbonate transporters are expressed in the CPE. In the basolateral membrane the electroneutral Na(+) dependent Cl(-)/HCO3 (-) exchanger, NCBE (slc4a10) is expressed. In the luminal membrane, the electrogenic Na(+):HCO3 (-) cotransporter, NBCe2 (slc4a5) is expressed. The electroneutral Na(+):HCO3 (-) cotransporter, NBCn1 (slc4a7), has been located in both membranes. In addition to the bicarbonate transporters, the Na(+)/H(+) exchanger, NHE1 (slc9a1), is located in the luminal membrane of the CPE. Genetically modified mice targeting slc4a2, slc4a5, slc4a7, slc4a10, and slc9a1 have been generated. Deletion of slc4a5, 7 or 10, or slc9a1 has numerous impacts on CP function and structure in these mice. Removal of the transporters affects brain ventricle size (slc4a5 and slc4a10) and intracellular pH regulation (slc4a7 and slc4a10). In some instances, removal of the proteins from the CPE (slc4a5, 7, and 10) causes changes in abundance and localization of non-target transporters known to be involved in pH regulation and CSF secretion. The focus of this review is to combine the insights gathered from these knockout mice to highlight the impact of slc4 gene deletion on the CSF production and intracellular pH regulation resulting from the deletion of slc4a5, 7 and 10, and slc9a1. Furthermore, the review contains a comparison of the described human mutations of these genes to the findings in the knockout studies. Finally, the future perspective of utilizing these proteins as potential targets for the treatment of CSF disorders will be discussed.