Frontal White Matter Biochemical Abnormalities in Late-Life Major Depression Detected With Proton Magnetic Resonance Spectroscopy

Mood disorders are among the most common mental disorders in the elderly population (1). Both major depressive disorder, the most severe form of depression, and clinically significant forms of minor depression are responsible for considerable psychosocial morbidity in elderly patients (2). These effects include frequent visits to physicians’ offices and emergency rooms, abuse of alcohol and tranquilizers, absenteeism, and suicide (2). Mood disorders in elderly patients are also frequently associated with a broad spectrum of medical disorders, including cardiovascular, cerebrovascular, pulmonary, and musculoskeletal diseases (3, 4). Despite these consistent epidemiological and clinical observations, the precise biological underpinnings of depression in late life remain largely unknown.

Both physiological and neuroanatomical approaches have been used to examine the neurobiological correlates of depression in late life (5–11). Studies with positron emission tomography and xenon-133 inhalation methods have shown lower levels of glucose metabolism and cerebral blood flow in neocortical and subcortical regions in elderly patients with major depression than in comparison subjects (5, 6). Magnetic resonance imaging (MRI) studies have revealed two kinds of structural abnormalities in patients with late-life depression: lower brain volumes in circumscribed regions and a higher volume of high-intensity lesions in the parenchyma—areas that appear bright on T₂-weighted MRI findings (7–11). The lower brain volumes are more striking in the prefrontal lobes, hippocampus, and head of the caudate nucleus (9). High-intensity lesions occur predominantly in the periventricular and deep white matter regions, although they have also been described in the subcortical nuclei (10).

The frontal lobes have been frequently implicated in the pathophysiology of psychiatric disorders (12–14). By virtue of their rich afferent and efferent connections with other neocortical, limbic, and subcortical regions, they play an important role in the regulation and modulation of affect and emotions (12–14). MRI high-intensity lesions are widely considered to provide some of the neurobiological substrates of depression and occur largely in the white matter regions of the brain. Despite these observations, to our knowledge there are no published reports on the biochemical changes in the gray and white matter regions of the frontal lobe in patients with late-life major depression.

The purpose of this study was to examine the levels of N-acetylaspartate, choline, myo-inositol, and creatine in the left dorsolateral white matter of the frontal lobe and bilaterally in the anterior cingulate gray matter of patients with late-life major depression and a healthy comparison group. We hypothesized that levels of N-acetylaspartate would be lower and levels of choline would be higher in the frontal white and gray matter in patients with major depression than in the comparison subjects. These hypotheses were based on our prior observations and on published reports of anatomical and physiological abnormalities in the frontal lobes in patients with mood disorders.

Method

Subjects

The study groups consisted of 20 patients with late-life major depression (14 women, six men, mean age=69.95 years, SD=8.18) and 18 nondepressed comparison subjects (13 women, five men, mean age=72.39 years, SD=5.79). All depressed patients were age 60 or older and met DSM-IV criteria for major depressive disorder. All patients had scores of 15 or greater on the 17-item Hamilton Depression Rating Scale (15). In addition to receiving a detailed mental status examination by a psychiatrist, all patients were assessed with a structured psychiatric interview (Structured Clinical Interview for DSM-IV). Patients and comparison subjects were free of psychotropic medications for at least 2 weeks before the scan. Patients received comprehensive medical and neurologic examinations and laboratory tests, including complete and differential blood counts; liver, renal and thyroid screening tests; and measurement of electrolyte levels. None of the patients and comparison subjects had mental status examinations consistent with clinical dementia or any other brain disorder. Their mean Mini-Mental State Examination (16) scores were in the normal range (mean=28.45, SD=1.54, in the major depression group, and mean=29.53, SD=0.80, in the comparison group). Patients were recruited through local newspapers and radio advertisements and through referrals from the geriatric psychiatry ambulatory care programs at the UCLA Medical Center. Comparison subjects were recruited from the community through newspaper and radio advertisements. All aspects of the study were explained to the subjects before the study started, and informed consent was obtained in keeping with the guidelines of UCLA’s human subjects protection committee.

Single-Voxel Proton Spectroscopy Methods

Acquisition

Water-suppressed hydrogen 1 (¹H) magnetic resonance (MR) spectra were acquired in the anterior cingulate gray and the left dorsolateral prefrontal white matter regions. A voxel size of 8 ml was localized by using the PRESS sequence and CHESS for water suppression (P. Bottomley, U.S. Patent 4-480-228.1984) (17, 18). Figure 1 shows the voxel locations in an MRI scan of the brain of a comparison subject. Numerically optimized Shinnar-Le Roux radiofrequency pulses were used for PRESS (90°, 180°, 180°) and CHESS (90°, 90°, 90°).

The following parameters were used for MR spectral acquisition: TR=3 seconds, TE=30 msec, and NEX=64. Four unsuppressed water free-induction delays were also acquired from each location for eddy current compensation and phase correction (19). A GE 1.5-T MRI/magnetic resonance spectroscopy (MRS) scanner (General Electric Medical Systems, Waukesha, Wis.) operating in the 5.8 mode with echo-speed gradients (maximum 22 mT/m) was used. A standard birdcage quadrature MRI head coil was used for transmission/reception. The MRS protocol consisted of acquisition of metabolite and water free-induction delays after optimization of 90°/180° radiofrequency pulses, automated shimming using the localized water signal and water suppression.

MRS post-processing

The MRS raw files were transferred to an SGI off-line O2 workstation (Silicon Graphics Inc., Sunnyvale, Calif.). The raw files were processed by using the LC-Model algorithm (20). The water-suppressed time domain data were analyzed between 1.0 ppm and 4.0 ppm without further T₁ and T₂ correction (Figure 2). The basis set provided by the vendor (20) was used and then scaled to a consistent transmitter gain. Absolute values of N-acetylaspartate, creatine, choline, and myo-inositol are reported in mM uncorrected for T₁ and T₂ saturation. The absolute metabolite values were further corrected for CSF by segmenting the three-dimensional spoiled gradient recalled echo coronal MR images into gray matter, white matter, and CSF.

Anatomical landmarks

The axial slice showing the most anterior extent of the anterior margin of the genu of the corpus callosum was chosen as the reference image on which to center the voxels for both locations. This was determined by placing a cursor at the anterior margin of the genu in the midline, on several axial slices through the genu. The slice showing the most anterior location of the anterior margin of the genu was then chosen as the reference slice. This anatomical landmark was chosen because it is an identifiable single location, which helps to standardize voxel placement despite variations in scanning angle. Other potential single location sites in the brain are at a greater distance from the areas of interest, and, therefore, the potential error related to variation in scanning angle is greater.

For anterior cingulate gray matter, the posterior boundary of the voxel was placed on the reference slice immediately adjacent to the anterior margin of the genu of the corpus callosum. The cingulate gyrus lies immediately anterior to the anterior margin of the genu of the corpus callosum. The voxel lies symmetrically across the interhemispheric fissure. The area covered by the voxel corresponds to the rostral cingulate region, partially occupying rostral Brodmann’s areas 24 and 32. This region has been implicated in the regulation and modulation of affect and cognition. For the dorsolateral prefrontal cortex, the medial border of the voxel was placed contiguous to the gray-white juncture on the medial side of the hemisphere by using the reference slice described earlier. The posterior boundary of the voxel was placed immediately adjacent to the anterior margin of the frontal horn of the lateral ventricle.

MRI methods

All images were taken at 1.5 T Signa scanner (General Electric Medical Systems, Milwaukee) by using a standardized imaging protocol. Measurements of intracranial gray matter, white matter, and CSF volumes were derived from a T₁-weighted, three-dimensional volumetric spoiled gradient recalled echo sequence with 124 contiguous partitions, 1.6-mm slice thickness, 22×16.5 cm field of view, 192 views, and 45° flip angle. The spectroscopy voxels (2×2×2 cm³) were localized on fast spin echo images acquired at the same scanning session as the three-dimensional spoiled gradient recalled echo data; thus, when no head movement occurred between these scans, the two scans were in register and the location of the spectroscopy data could be easily mapped to the three-dimensional spoiled gradient recalled echo data. When subject movement occurred between the fast spin echo and three-dimensional spoiled gradient recalled echo data, a rigid body alignment (21) was performed on the three-dimensional data to correct for misalignment, allowing localization of the spectroscopy voxel within the three-dimensional data.

Segmentation of the three-dimensional spoiled gradient recalled echo data was accomplished with a minimum distance classifier algorithm, which was used previously for this T₁-weighted imaging protocol with good results in our laboratory (22). After brain masks were created, a three-dimensional spatial filter was applied to each data set to remove low-frequency signal drifts across the image data (23). The filtered images were resubmitted to the minimum distance tissue classifier, thus producing gray matter, white matter, and CSF trinary tissue counts for total brain voxels. Each spectroscopy voxel was then localized to the subject’s segmented tissue map to derive the volume (mm³) of gray matter, white matter, and CSF that made up the spectroscopy voxel, permitting partial volume correction of the spectroscopy data.

Statistical Analysis

Both mean absolute levels (mM) and mean ratios of metabolite to creatine levels were compared across groups. Analysis of covariance (ANCOVA), controlling for age, gender, and possible age-by-gender interactions, was used to compare means between the patient and comparison groups. Ratios were obtained by using creatine level as the denominator because this pattern is widely reported in the literature and also because creatine levels were not significantly different between groups. ANCOVA, controlling for age, gender, and possible age-by-gender interactions, was also used to compare groups on the gray and white matter (as a percentage of brain volume) contents of the voxels.

Results

The choline/creatine and myo-inositol/creatine ratios were significantly higher in the dorsolateral white matter in the major depression group than in the comparison group (Table 1, Figure 3). There were no statistically significant differences in the N-acetylaspartate/creatine ratio between the two groups. None of the metabolite ratios in the gray matter differed between the patient and comparison groups (Table 1). There were no differences between groups in the absolute metabolite levels in either the gray or the white matter. Gray and white matter composition of the voxels was comparable in both groups (Table 2).

Discussion

To our knowledge, our study is the first to demonstrate that frontal lobe biochemical abnormalities in patients with late-life major depression are more striking in the white than in the gray matter. The biochemical abnormalities observed in our study consisted of more prominent choline/creatine and myo-inositol/creatine signals in the left dorsolateral white matter in patients with major depression than in the comparison subjects. There was no significant difference in the white matter N-acetylaspartate levels between patients and comparison subjects. Also, there were no significant differences in any of the major metabolites in the gray matter between the two groups. These data suggest that physiological abnormalities in the white matter may provide an important neurobiological substrate to depression, especially in elderly patients.

Although higher choline resonances have been demonstrated in younger patients with both unipolar and bipolar depression, these results are controversial (24–26). The bulk of the evidence appears to support higher choline resonances in younger patients, relative to comparison subjects, especially in the basal ganglia/temporal lobe region (24–26). Another report, however, demonstrated lower choline/creatine ratios in the basal ganglia/temporal lobe regions in patients, especially in nonresponders to pharmacological intervention, than in comparison subjects (27). The only published study of elderly patients that we could find (28) demonstrated a higher choline/creatine ratio in a large voxel that encompassed several subcortical structures in seven patients with major depression, relative to comparison subjects. In this study, the high choline/creatine ratios normalized with pharmacological treatment.

The choline resonance obtained from in vivo MRS has been shown to reflect free choline, phosphocholine, and glycerophosphocholine (29). It is therefore a composite signal from several choline-containing metabolic compounds (30). Choline is an essential precursor of acetylcholine and has been implicated in the pathophysiology of mood disorders. Indirect evidence from several pharmacological studies suggests that choline may have a “depressogenic” effect on the central nervous system (CNS) (31). It has been suggested that higher levels of choline resonance on MRS may reflect alterations in the structure of neuronal membranes of which choline components are an integral part. However, the precise relationship between the choline signal on MRS and acetylcholine transmission in the brain is unclear. The use of phosphorous (³¹P) spectroscopy in conjunction with high-field proton MRS is likely to provide additional information in this regard.

The myo-inositol signal in MRS is also likely to be a composite signal with the bulk of the contributions coming from myo-inositol itself (32, 33). The other two contributors to this signal are myo-inositol monophosphate and glycine. Although inositol is presumed to play an important role in cerebral metabolism, its precise role remains unclear. myo-Inositol may serve as the storage form of the inositol diphosphate that links receptors to intracellular activity, thereby serving as secondary or third messengers for hormones and neurotransmitters in the CNS (32). myo-Inositol is also the substrate for the mood stabilizing agent lithium, which reduces myo-inositol levels by inhibiting the enzyme inositol monophosphate, a catalyst for converting inositol monophosphate hydroxyls to myo-inositol. myo-Inositol/creatine levels in the parietal cortex decrease in response to neuronal insult in conditions like hepatic encephalopathy (33–35). Explanations for the observed higher levels of myo-inositol include higher levels of myo-inositol uptake or retention or changes in the cellular/extracellular matrix. Alternative explanations include a possible perturbation in the coupling metabolism of the receptor-secondary messenger system complex, thereby providing an important biological substrate to mood disorders in late life.

Several subtle and overlapping biological factors need to be considered in interpreting the changes in metabolite levels in the brain in specific clinical brain disorders (36–41). The levels of some of these metabolites in certain brain regions have been reported to be higher in elderly subjects, compared with younger subjects (37). There are also likely to be regional differences in the levels of metabolites in the CNS (36, 39, 41). Further, the effect of aging might attenuate regional metabolite concentrations in the brain. The underlying tissue composition, gray versus white matter, has also been shown to influence metabolite levels (38–41). Consequently, the anatomical location and the underlying tissue composition of the region examined are critical to the validity and biological significance of these measurements. Age- and gender-matched comparison subjects need to be included in studies examining biochemical changes associated with specific brain disorders.

Although lower volumes of focal brain regions have been described in patients with late-life depression, reductions in N-acetylaspartate are seldom seen in MRS studies of patients with significant mood disorders. N-Acetylaspartate is traditionally considered a marker of the structural integrity of the neuron, both of the cell body and the axon, and the level of N-acetylaspartate is typically low in degenerative disorders in which there is demonstrable cell loss (42, 43). Higher levels of choline and myo-inositol signals in the absence of changes in N-acetylaspartate raise the question of the role of the nonneuronal compartment in the pathophysiology of mood disorders. The focus of in vivo and in vitro studies in the realm of behavior and higher cortical functions has typically been the neuron. The role of glia in the etiology of clinical brain disorders in general and psychiatric illness in particular remains unknown. The glial compartment and its physiological role have been receiving increasing attention recently (44). The myo-inositol system together with acetylcholine, glutamate, and other neurotransmitters may play an important role in glial cell function and in modulating synaptic activity (44). Therefore, in vivo estimates of choline and myo-inositol may also reflect biological changes in nonneural tissue, with implications for neuronal and synaptic function. Our findings corroborate physiological studies that demonstrate abnormalities in glucose metabolism and perfusion in the frontal lobes in patients with late-life major depression (5, 6). More sophisticated in vivo and in vitro techniques need to be developed before definitive statements on the glial-axonal axis can be made.

Some limitations of our methodologic approach should be discussed. First, overlap of metabolites’ spectra is a major problem with one-dimensional MR spectroscopy. The spectra for glutamine, N-acetylaspartate, γ-aminobutyric acid, and aspartate overlap in the 2–3 ppm region (45, 46). Further, myo-inositol, glutamate/glutamine, glucose, and aspartate spectral peaks overlap in the 3.5–4 ppm region. Time-domain fitting with the LC Model is more accurate for quantitation of overlapping peaks than conventional frequency domain fitting with the Marquardt algorithm. Despite this improvement, one-dimensional acquisition has intrinsic limitations that preclude a more precise characterization of brain metabolites. Localized two-dimensional correlated spectroscopy, especially if it is adapted to high-field systems, is expected to improve the analysis of several overlapping metabolites (46). Second, although voxel placement was designed to maximize tissue homogeneity, the relatively large size of our voxel (2×2×2 cm³) resulted in inclusion of both gray and white matter in each voxel location. However, since the voxel placed in the dorsolateral region comprised predominantly white matter, we feel confident that our findings primarily reflect biochemical changes in the frontal white matter. Smaller voxels obtained through chemical-shift imaging will allow more precise measurement of metabolite levels in circumscribed brain regions.

In conclusion, our observations suggest that some of the biochemical correlates of late-life major depression occur in the white matter of the frontal lobes. In general, the role of the white matter in most psychiatric diseases is underappreciated. The white matter has important roles in biological processes such as axoplasmic transport and neurotransmitter release. Disruption of these processes could lead to a cascade of changes that form the substrates of a clinical brain disorder. More focused approaches examining anatomical and biochemical changes in the white matter, both antemortem and postmortem, are likely to reveal the broad spectrum of changes that characterize mood and other psychiatric disorders.

TABLE 1

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TABLE 2

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