Glial dysfunction in abstinent methamphetamine abusers

Abstract

Persistent neurochemical abnormalities in frontal brain structures are believed to result from methamphetamine use. We developed a localized (13)C magnetic resonance spectroscopy (MRS) assay on a conventional MR scanner, to quantify selectively glial metabolic flux rate in frontal brain of normal subjects and a cohort of recovering abstinent methamphetamine abusers. Steady-state bicarbonate concentrations were similar, between 11 and 15 mmol/L in mixed gray-white matter of frontal brain of normal volunteers and recovering methamphetamine-abusing subjects (P>0.1). However, glial (13)C-bicarbonate production rate from [1-(13)C]acetate, equating with glial tricarboxylic acid (TCA) cycle rate, was significantly reduced in frontal brain of abstinent methamphetamine-addicted women (methamphetamine 0.04 micromol/g per min (N=5) versus controls 0.11 micromol/g per min (N=5), P=0.001). This is equivalent to 36% of the normal glial TCA cycle rate. Severe reduction in glial TCA cycle rate that normally comprises 10% of total cerebral metabolic rate may impact operation of the neuronal glial glutamate cycle and result in accumulation of frontal brain glutamate, as observed in these recovering methamphetamine abusers. Although these are the first studies to define directly an abnormality in glial metabolism in human methamphetamine abuse, sequential studies using analogous (13)C MRS methods may determine 'cause and effect' between glial failure and neuronal injury.

Figure 1

Glial versus neuronal metabolism: pathway of ¹³C label from [1-¹³C]acetate to tricarboxylic acid (TCA) cycle in human brain. Selective metabolism of acetate and glucose occurs in glia and neurons, respectively. Only the relevant intermediates are shown. Glia selectively transports acetate which is then metabolized through acetate thiokinase and reactions of the mitochondrial tricarboxylic acid (TCA) cycle forming ¹³C glutamate and glutamine, enriched in the C1 and C5 positions, before final oxidation and release as ¹³C bicarbonate (CO₂=HCO₃⁻). Five of the relevant cerebral metabolic products are showed in sequential ¹³C brain spectra of Figure 3. The alternative metabolic pathways of ¹³C glucose metabolism, predominantly in neurons, but not contributing to this study in which only the single substrate, [1-¹³C]acetate is provided. Ac, acetate; Glc, glucose; Glu, glutamate; Gln, glutamine; CO₂, carbon dioxide; Pyr, pyruvate; V_g glc, glial glucose oxidation; V_g ac, glial acetate oxidation; α-KG, α-ketoglutarate; V_glu, glutamate synthesis rate; V_gln, glutamine synthesis rate. Glucose is metabolized in neuron and glia whereas acetate is metabolized exclusively in glia.

Figure 2

Human ¹³C MR frontal brain spectra acquired during [1-¹³C]acetate infusion: carbon-13 magnetic resonance spectroscopy (MRS) at baseline and 120 mins after the infusion (154 to 164 p.p.m.) in a control and an abstinent methamphetamine-dependent (AMD) subject. ¹³C MRS spectra show HCO₃⁻ and total Cr (tCr) chemical shift region (154 to 164 p.p.m.) at baseline and at the end of the infusion study in a normal control subject (A) and an AMD subject (B). The lower, natural abundance spectra for the normal control (left) indicate that cerebral bicarbonate concentration is very similar to that of [Cr+PCr], i.e., 11 mmol/L. The resting cerebral bicarbonate concentration in a representative AMD subject (right) was indistinguishable from the control (HCO₃=11 mmol/L). After 2 h of [1-¹³C]acetate infusion, substantial but different increases in cerebral ¹³C bicarbonate intensity were observed. Again, by comparing the adjacent, unenriched Cr+PCr, the rate of enrichment of cerebral bicarbonate was substantially lower in the AMD (right) than in the normal control (left). These representative data are seen to be statistically significantly different when normal and AMA subjects are compared in Tables 1 and 2. In the control subject, the concentrations of bicarbonate determined in this way were 15 μmol/g at baseline and 49 μmol/g at the end of the 2 h infusion study (Cr does not become enriched) whereas in the AMD subject initial bicarbonate was 10 μmol/g at baseline but reached only 25 μmol/g after 2 h.

Figure 3

Time courses of glutamate C5 and acetate (A), glutamine C5 (B), and HCO₃⁻ (C) from a healthy subject. Signal intensities are shown in arbitrary units. Note that glutamine C5, glutamate C5, and bicarbonate become increasingly enriched over time, consistent with the expected path of glial metabolism of ¹³C acetate shown in Figure 1. After significant enrichment up to 60 to 80 mins, pools of glutamate C5 and glutamine decline, presumably as they are in turn metabolized to the final product, ¹³C HCO₃⁻. No significant sequestration in glutamate and glutamine occurs.

Figure 4

Effect of methamphetamine abuse on cerebral [1-¹³C]acetate metabolism. Frontal brain ¹³C metabolites observed during [1-¹³C]acetate infusion: figure shows carbon-13 MRS (150 to 195 p.p.m.) in a control and an abstinent methamphetamine-dependent subject after enrichment with [1-¹³C]acetate. Stacked sequential carbon-13 spectra and spectra summed for the entire infusion protocol (120 mins, bottom) are shown for a control (A, B) and an abstinent methamphetamine-dependent (AMD) subject (C, D). The spectra identified enriched cerebral metabolites glutamate C5+C1 acetate (182 p.p.m.), glutamine C5 (178 p.p.m.), glutamate C1+glutamine C1 (175 p.p.m.), and the final oxidation product, bicarbonate (161 p.p.m.).

Figure 5

Effect of methamphetamine abuse on cerebral metabolism of ¹³C acetate. Time courses of appearance of the end product of [1-¹³C]acetate oxidation, HOC₃⁻ are compared. Low-power noise nuclear overhauser effects (NOE) spectra were acquired in 6.5 min blocks for up to 120 mins after enrichment by i.v. acetate (3 mg/kg body weight) from frontal brain in five abstinent methamphetamine-dependent (AMD) subjects and five control subjects. Logarithmic fits of the averaged percent fractional enrichment of bicarbonate (%E) were plotted as a function of time after start of the infusion. %E in AMD (lower trace) is shown to be lower than controls (upper trace). Standard deviations are also shown as cross-bars.

Figure 6

Comparison of methods to express effect of abstinent methamphetamine-dependent (AMD) in ¹³C HCO₃⁻ production from [1-¹³C]acetate in human frontal brain. Results shown in Figure 5 are quantified. Means and standard deviation of percent fractional bicarbonate enrichment calculated at three different time points (60, 80, and 120 mins) and the initial slope of bicarbonate production from control and AMD groups. P-value in each group comparison is <0.001.