Y.L. Yamamoto

A new method to measure brain serotonin synthesis in vivo. I. Theory and basic data for a biological model.

We describe here an autoradiographic method to measure the in vivo rate of serotonin synthesis in rat brain. The method is based on the use of the l-tryptophan analogue a-methyl-l-tryptophan (a-MTrp), which is converted in vivo into a-methylserotonin (a-M5HT). Since a-M5HT is not a substrate for monoamine oxidase, it is accumulated in the brain tissue. Data are presented to confirm time-dependent conversion of a-MTrp into a-M5HT in the dorsal raphe nucleus and also in the pineal body, an organ outside the blood–brain barrier. It has also been shown that washing brain slices in 10% trichloroacetic acid results in <3% incorporation of a-MTrp into brain proteins. The rates of synthesis are calculated in several grossly dissected brain structures by using tracer kinetics and a three-compartment biological model. The half-life of the precursor pool is estimated to be ∼20 min. The rate of serotonin synthesis is highest in the pineal body.

Serotonin synthesis rate measured in living dog brain by positron emission tomography

Abstract: In vivo measurements by positron emission tomography of the brain serotonin synthesis rates in the normal dog, in the dog with increased plasma tryptophan concentration, and in the dog under different arterial oxygen tensions are described. The method described here permits repeated measurements in the same brain for the first time. An increase in the plasma tryptophan concentration from 16.6 to 191.5 and then to 381 μM resulted in close to a linear increase in the brain serotonin synthesis rate. When PaO2 was raised from 76 ± 2 to 106 ± 1 mm Hg, the rate of serotonin synthesis in the dog brain increased from 39 ± 8 to 54 ± 10 pmol g−1 min−1. The estimates of the Michaelis‐Menten constants, Kappm and Vmax for the transport of tryptophan through the blood‐brain barrier are 303 ± 54 μM and 63 ± 10 nmol g−1, min−1, respectively.

A new method to measure brain serotonin synthesis in vivo. II. A practical autoradiographic method tested in normal and lithium-treated rats.

We describe here a practical autoradiographic method to estimate the rate of serotonin synthesis in brain. A two-time point method (60 and 150 min after injection of a-[14C]methyl-l-tryptophan) was first evaluated in 14 normal rats (7 at each time point). After this the method was tested in lithium-treated rats. In normal rats the rate of serotonin synthesis measured by the two-time point method generally correlated with known concentrations of tryptophan hydroxylase. The rate of synthesis in lithium-treated rats was compared with that in shamtreated rats (NaCl treatment). The results showed a significant increase in the synthesis rate in some cerebral structures. The greatest increases in the serotonin synthesis rate, attributable to the lithium treatment, were observed in the parietal cortex (52%) and caudate nucleus (47%). This is the first investigation to demonstrate, with autoradiographic resolution (∼100 μm), the differential changes in the rate of serotonin synthesis in the brain. Lithium had no significant effect on the rate of synthesis in the pineal gland.

Autoradiographic determination of brain pH following middle cerebral artery occlusion in the rat.

A method of quantitative autoradiography usingMC-labelled 5,5-dimethyl-2,4-oxazolidin- edioneI4C-DMO to evaluate the local changes in brain pH after ischemia is described. In normal control raU the calculated tissue pH values in gray matter were slightly lower than those in white matter, and there was no significant difference in the calculated pH among the various structures in cortical and subcortical gray matter. Four hours after a left middle cerebral artery (MCA) occlusion, marked reductions in14C- DMO concentrations were demonstrated in the anterior two-thirds of the cerebral cortex and in the lateral part of the caudate nucleus indicating tissue acidosis in these areas. Although several assumptions are required for the calculation of pH in brain tissue, this method would appear very useful in the investigation of the altered metabolic state In ischemlc brain. Stroke VOL 15, No 3, 1984

Effect of Vascular Activity in the Determination of Rate Constants for the Uptake of 18F-Labeled 2-Fluoro-2-Deoxy-D-Glucose: Error Analysis and Normal Values in Older Subjects

Regional cerebral blood volume (CBV) can be calculated using data obtained during the kinetic analysis of 18F-labeled 2-fluoro-2-deoxy-d-glucose (FDG) uptake measured by positron emission tomography (PET). As a result the influence of vascular activity upon the determination of FDG rate constants can be minimized. The method is investigated by simulation experiments and by analysis of PET studies on seven older, healthy human volunteers aged 52–70 years. The accuracy of measured FDG rate constants k1, k2, and k3, obtained either by omitting the early portion of the uptake curve or by explicit inclusion of CBV as a fit parameter, is compared. The root mean square error in measured rate constant for the latter method is equivalent to that obtained by omitting the first 2.5–3 min of tissue data and neglecting the CBV term. Hence, added information about the physiological state of the tissue is obtained without compromising the accuracy of the (FDG) rate constant measurement. In hyperemic tissue the explicit determination of the vascular fraction results in more accurate estimates of the FDG rate constants. The ratio of CBV determined by this method to CBV obtained using C15O in six subjects with CBV in the normal range was 0.92 ± 0.32. A comparison of the CBV image obtained by this method with that obtained using C15O in an arteriovenous malformation case demonstrates the accuracy of the approach over a wide range of CBV values. The mean value for CBV fraction in gray matter obtained by this method in the older control group was 0.040 ± 0.014. Average gray matter rate constants obtained were k1 = 0.084 ± 0.012, k2 = 0.150 ± 0.071, and k3 = 0.099 ± 0.045 min−1.

Correlation of local cerebral blood flow, glucose utilization, and tissue pH following a middle cerebral artery occlusion in the rat.

The use of three sets of the double-tracer autoradiographic technique to measure topographical changes of local cerebral blood flow (LCBF), glucose utilization (LCGU), and tissue pH following a 3 h middle cerebral artery (MCA) occlusion in the rat is described. In a sham-operated group of animals there was 10% reduction of LCBF and 7% reduction of LCGU in the most affected areas as compared to the contralateral homologous regions. However, the ratio of LCGU/LCBF in the affected areas remained within normal limits. In the MCA-occluded animals, LCGU showed a bimodal response to decreased LCBF. LCGU decreased with reduced LCBF until LCBF fell to 38% of normal. Below this LCBF level LCGU increased, most likely implying anerobic glycolysis. Decline of tissue pH corresponds to the mismatch of LCBF and LCGU. These results suggest that brain tissue pH change cannot be predicted on the basis of LCBF or LCGU alone.

Positome II: A High Efficiency Positron Imaging Device for Dynamic Brain Studies

The performance of a new positron imaging device designed for high resolution dynamic studies on the human brain is described. This unit is the first to exploit the high photoelectric efficiency of bismuth germanate (BGO) detectors which are almost 3 times as dense as NaI. In the first 4 months of clinical use some 300 patient studies have been performed. The detector performance and design criteria are discussed. The principles of operation of the coincidence circuit and computer programs are described. Finally some clinical studies are presented to demonstrate its imaging capabilities.

Triple-tracer autoradiography demonstrates effects of hyperglycemia on cerebral blood flow, pH, and glucose utilization in cerebral ischemia of rats.

Triple-tracer autoradiography was used to measure topographic changes in local cerebral blood flow, cerebral tissue pH, and local cerebral glucose utilization in hyperglycemic and normoglycemic rats, all of which had undergone occlusion of the middle cerebral artery. More severe and extensive reduction of all three variables was observed in the hyperglycemic than in the normoglycemic rats. In seven normoglycemic rats, significant reduction in local cerebral blood flow (p less than 0.025) was observed in the ischemic but not in the contralateral nonischemic side at the lateral portion of the caudate nucleus and the neocortex. Tissue pH was significantly lower (p less than 0.025) only at the lateral portion of the caudate nucleus in the ischemic side. No significant differences in local cerebral glucose utilization were observed when the two hemispheres were compared. In the ischemic hemisphere of five hyperglycemic rats, the caudate nucleus and the neocortex exhibited significant reduction (p less than 0.025) in local cerebral blood flow, tissue pH, and local cerebral glucose utilization. Even in the nonischemic hemisphere of the hyperglycemic rats, local cerebral blood flow in the caudate nucleus and the neocortex was significantly reduced (p less than 0.025) compared with the normoglycemic rats. No significant change in tissue pH or local cerebral glucose utilization was observed throughout the nonischemic hemisphere of the hyperglycemic compared with the normoglycemic rats. Tissue pH was systematically lower in the hyperglycemic than in the normoglycemic rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Hemodynamic and metabolic effects of cerebral arteriovenous malformations studied by positron emission tomography

Seventeen patients with an intracranial arteriovenous malformation were studied with positron emission tomography. Cerebral blood flow, cerebral blood volume, oxygen extraction fraction, and glucose and oxygen metabolism were evaluated in both hemispheres, excluding the area of the malformation itself. Patients were divided into three groups according to the size of their malformation, and results obtained were compared with studies in healthy volunteers. The glucose metabolism was significantly (p less than 0.01) decreased in the ipsilateral hemisphere in all patients. The cerebral blood volume was significantly increased (p less than 0.001) ipsilaterally in the three groups, and contralaterally in patients with medium- and large-sized arteriovenous malformations. The cerebral blood volume to cerebral blood flow ration, an index of vascular mean transit time, was significantly increased (p less than 0.005) ipsilaterally in patients with medium- and large-sized malformations and contralaterally in patients with large ones. Cerebral blood flow, oxygen extraction fraction, and oxygen metabolism were within the normal range bilaterally in all three groups, but oxygen extraction fraction tended to be higher in patients with larger lesions. The lack of significant change in oxygen metabolism suggests that oxygen metabolism in cortical areas remote from the arteriovenous malformation has been maintained by compensatory hemodynamic mechanisms. These data reveal widespread metabolic and hemodynamic consequences of arteriovenous malformations and suggest that they are associated with impairment of glucose metabolism, both in ipsilateral regions remote from the lesion and in the contralateral hemisphere in patients with large lesions.

Rapid steady-state analysis of blood-brain transfer of l-Trp in rat, with special reference to the plasma protein binding

We estimated constants for the binding of tryptophan (Trp) to plasma proteins, and for the transfer of Trp from plasma to brain in rat. The measurements were made under conditions in which the plasma and brain concentrations of Trp were raised to new steady-states for at least 10 min before being measured. The concentration of other competing amino acids were also at a steady-state. The plasma Trp concentration was elevated by i.p. injection of different doses of L-tryptophan methyl ester 60 min before the measurement of the plasma-brain transfer. We simultaneously measured blood flow with [14C]-butanol, and the brain tissue Trp uptake with [3H]Trp. The maximal velocity (Vmax), apparent half-saturation Michaelis-Menten constant (Km(app)), and diffusion constant (PdS) for Trp transport from plasma into brain were found to be 7.0 +/- 2.1 nmol g-1 min-1, 36 +/- 17 microM, and 0.065 +/- 0.006 ml g-1 min-1, respectively. The maximum plasma protein binding (Bmax) and dissociation constant (KD) for Trp were estimated at 360 +/- 16 nmol/ml-plasma and 81 +/- 10 microM, respectively. We conclude that the plasma protein binding of Trp inhibits the blood-brain transfer in inverse proportion to the plasma free Trp concentration.