We determined cerebral intracellular pH in living rabbits and rats under physiologic conditions, using phosphorus NMR spectroscopy and new titration curves thought to be appropriate for brain. Mean values for the two species were, respectively, 7.14 +/- 0.04 (SD) and 7.13 +/- 0.03. These are toward the alkaline end of the range of values obtained by other methods, as values reported by other NMR workers also tend to be.

1H NMR spectra at 360.13 MHz were obtained of the rat brain in vivo by using a surface coil placed over the skull. Resonances of numerous metabolites were identified by comparison with the 1H NMR spectra of excised rat brain tissue and acid extracts of the tissue. Changes in cerebral lactate levels resulting from the administration of gas mixtures low in oxygen were monitored in the in vivo brain spectra with a time resolution of 2.3 min. The electroencephalogram and electrocardiogram were recorded simultaneously during the NMR experiment. Reversibility of the hypoxic response was documented when, upon oxygen administration, cerebral lactate returned to its prehypoxic level. These experiments demonstrate the applicability of high-resolution 1H NMR to monitor pathophysiology of brain metabolism in real time.Nuclear magnetic resonance spectroscopy (NMR) is rapidly developing into a major tool for the study of metabolism in vivo. Both 31p and 13C NMR have been used extensively in metabolic studies of cellular suspensions and perfused organs (1, 2). The recent introduction of surface coils (3) has led to the successful application of topical 31P NMR to studies of the vital organs of animals and man (4-6). Recently, topical 13C NMR has been shown to be capable of monitoring glycogen stores and acyl glycerides in vivo (7-9), greatly extending the kinds of metabolic information that can be obtained. MATERIALS AND METHODS Animal Preparation for in Vivo 1H Experiments. Rats of theCharles River strain (220-240 g), fed ad lib, were anesthetized with halothane (1%), tracheotomized, mechanically ventilated (25% 02/75% N20), and paralyzed with D-tubocurarine chloride (1.5 mg/kg, subcutaneously) and pancuronium bromide (1 mg/kg, subcutaneously). The animals were equipped with electrocardiogram leads on the left extremities and scalp leads for electroencephalogram recording. An incision was made along the midline of the calvarium so that skin and temporalis muscles-detached from the temporal ridge-could be retracted and fixed in position away from the skull. The rat was mounted vertically in a Plexiglass retainer and a small surface coil was centered 6 mm occipital to the bregma. These precautions ensure that only the contents of the skull are observed.Extract Preparation for 1H NMR. Extracts were prepared from rat brain frozen in situ (18) in the following manner: frozen cortical tissue was chipped away (68 mg) and extracted with 99% MeOH/0. 1 M HCl (2:1, wt/vol) and 0.1 M perchloric acid (10:1, wt/vol) according to well-established methods (19) with minor modifications. In the extraction procedure, 0.1 M sodium phosphate (pH 7.2) was substituted for imidazole buffer (19). After neutralization with 1.5 M KOH/0.3 M KCI the sample was lyophilized and dissolved in 0.38 ml of a 0.1 M phosphate/2H20 buffer (pH 7.2) and placed in a 5-mm NMR tube for analysis.Chemical shifts for resonances in excised and in vivo brain tissue were referenced to N-acetylaspartate at 2.023 ppm relative to sodium 3-trimethylsilyl[2,2,3,3-2H]pro...

We have used (13C)-1H NMR spectroscopy at 360.13 MHz to resolve the 13C coupled proton resonance of glutamate and lactate in the rat brain in vivo. The time required for the 13C fractional enrichment of the 4-CH2 position of brain glutamate to reach isotopic steady state was determined during a continuous infusion of D-[1-13C]glucose. Under conditions of ischemia, measurements made of the 3-CH3 of lactate in (13C)-1H NMR spectra revealed the relative contribution of brain glucose and glycogen to lactate formation. (13C)-1H NMR was 11 times more sensitive than 13C NMR for the detection of 13C in the 3-CH3 position of lactate and 6 times more sensitive for the detection of 13C in the 4-CH2 of glutamate under similar in vivo conditions.

Background-Neuroimaging and electrophysiological studies have consistently provided evidence of impairment in anterior cingulate cortex (ACC)/medial frontal cortex (MFC) function in people with schizophrenia. In this study, we sought to clarify the nature of this abnormality by combining proton magnetic resonance spectroscopy ( 1 H-MRS) with functional magnetic resonance imaging (fMRI) at 3T.

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We have analyzed changes in intracellular pH and phosphate metabolism during the cell cycle of Saccharomyces cerevisiae (NCYC 239) by using high-resolution 31P NMR spectroscopy. High-density yeast cultures (2 x 108 cells per ml) were arrested prior to "start" by sequential glucose deprivation, after which they synchronously replicated DNA and divided after a final glucose feeding. Oxygenation of arrested cultures in the absence ofglucose led to increased levels ofsugar phosphates and ATP and an increase in intracellular pH. However, these conditions did not initiate cell cycle progression, indicating that energization is not used as an intracellular signal for initiation ofthe cell division cycle and that the cells need exogenous carbon sources for growth. Glucose refeeding initiated an alkaline intracellular pH transient only in the synchronous cultures, showing that increased intracellular pH accompanies the traversal of start. Changes in phosphate flow and utilization also were observed in the synchronous cultures. In particular, there was increased consumption of external phosphate during DNA synthesis. When external phosphate levels were low, the cells consumed their internal polyphosphate stores. This shows that, under these conditions, polyphosphate acts as a phosphate supply. 31P NMR spectroscopy is rapidly gaining in importance as a technique for the noninvasive analysis ofphosphorus-containing metabolites and intracellular pH in vivo. Since the first 31p NMR spectra ofyeast were published in 1975 (1), sensitivity and resolution have been improved to allow for the investigation of glycolytic mutants (2), the control of glycolysis (3), and compartmentation of pH in ascospores (4). In this communication, we present the results of a 31P NMR analysis of synchronous suspensions of yeast cells in which we have investigated intracellular pH and phosphorus metabolism during the cell division cycle.Yeast are an ideal system for investigating the cell division cycle. We have found that they can be maintained for long periods at the high densities (2 x 108 cells per ml) necessary to obtain good signal-to-noise in the NMR experiments. In addition, many temperature-sensitive cell division cycle mutants of Saccharomyces cerevisiae are available, allowing for more detailed analyses in the future.Concepts of the cell division cycle in yeast have changed in recent years. Using the cell division cycle mutant system, Hartwell and his colleagues (5-10) have identified the presence of two loosely coupled subcycles, each comprising a series of discrete, interdependent steps. One subcycle involves the nuclear events of DNA replication and nuclear division; the other consists of the cytoplasmic events governing bud emergence and growth. The subcycles converge prior to mitosis and diverge again after a point called "start"; this point bears some similarity to the R point ofmammalian cells (11). Although much is known with regard to the morphologic and genetic aspects of "start," its biochemistry is relatively unknown. ...

The limited chemical shift dispersion of in uiuo "P NMR spectra obtained at the relatively low field strengths used for human applications is the cause of poor spectral resolution. This makes it di8fidt to obtain accurate quantitative information from overlapping resonances, and interesting resonances may be obscured. At 1.5 T unresolved 'H-3'P couplings contribute significantly to the liewidth of in uiuo "P NMR resonances. Therefore, proton decoupliig can improve spectral resolution substantially, resulting in better resolved resonances and more reliable quantitative information. In this work it is shown that well resolved resonances of glycerophosphocholine, glycerophosphoethanolamine and phosphoethanolamine are obtained in ' H decoupled "P N M R spectra of human muscle, brain, and liver. In spectra of the human heart it has been possible to resolve the myocardial Pi signal from the signals of 2,3-diphosphoglycerate from blood. With surface coils it is dficult to achieve broadband decoupliig over the entire sensitive region of the coil by using conventional decoupling sequences. This problem has been overcome by applying a train of frequency modulated inversion pulses to achieve proper decoupliig despite B, inhomogeneity. Broadband 'H decoupling of "P NMR spectra was possible without exceeding specific absorption rate guidelines.

Hydrogen-1 magnetic resonance (MR) spectroscopic images of patients with intracranial tumors were obtained. Metabolite maps of N-acetyl aspartate, choline, lactate, and creatine concentrations were reconstructed with a nominal spatial resolution of 7 mm and a section thickness of 25 mm. The metabolite maps showed variations in metabolite concentrations across the tumor. In one patient, it was observed that choline concentration was increased in one part of the tumor but decreased in another part. In another patient, the concentration of N-acetyl aspartate was extremely low in one part of the tumor but only slightly increased in another part of the tumor. Lactate was observed in all patients. In one patient, a combined measurement made with positron emission tomography (PET) and MR spectroscopic imaging was performed. This demonstrated that increased lactate concentration measured with H-1 MR spectroscopic imaging corresponded topographically with increased glucose uptake measured with fluorine-18 fluoro-2-deoxyglucose PET. Combined MR spectroscopic and PET measurements provide an opportunity to investigate, in greater detail than before, glucose uptake and catabolism by intracranial tumors.