Diffusion properties of major white matter tracts in young, typically developing children.

Brain development occurs rapidly during the first few years of life involving region-specific changes in both gray matter and white matter. Due to the inherent difficulties in acquiring magnetic resonance imaging data in young children, little is known about the properties of white matter in typically developing toddlers. In the context of an ongoing study of young children with autism spectrum disorder, we collected diffusion-weighted imaging data during natural nocturnal sleep in a sample of young (mean age=35months) typically developing male and female (n=41 and 25, respectively) children. Axial diffusivity, radial diffusivity, mean diffusivity and fractional anisotropy were measured at 99 points along the length of 18 major brain tracts. Influences of hemisphere, age, sex, and handedness were examined. We find that diffusion properties vary significantly along the length of the majority of tracks. We also identify hemispheric and sex differences in diffusion properties in several tracts. Finally, we find the relationship between age and diffusion parameters changes along the tract length illustrating variability in age-related white-matter development at the tract level.

Brain enlargement is associated with regression in preschool-age boys with autism spectrum disorders.

Autism is a heterogeneous disorder with multiple behavioral and biological phenotypes. Accelerated brain growth during early childhood is a well-established biological feature of autism. Onset pattern, i.e., early onset or regressive, is an intensely studied behavioral phenotype of autism. There is currently little known, however, about whether, or how, onset status maps onto the abnormal brain growth. We examined the relationship between total brain volume and onset status in a large sample of 2- to 4-y-old boys and girls with autism spectrum disorder (ASD) [n = 53, no regression (nREG); n = 61, regression (REG)] and a comparison group of age-matched typically developing controls (n = 66). We also examined retrospective head circumference measurements from birth through 18 mo of age. We found that abnormal brain enlargement was most commonly found in boys with regressive autism. Brain size in boys without regression did not differ from controls. Retrospective head circumference measurements indicate that head circumference in boys with regressive autism is normal at birth but diverges from the other groups around 4-6 mo of age. There were no differences in brain size in girls with autism (n = 22, ASD; n = 24, controls). These results suggest that there may be distinct neural phenotypes associated with different onsets of autism. For boys with regressive autism, divergence in brain size occurs well before loss of skills is commonly reported. Thus, rapid head growth may be a risk factor for regressive autism.

Associations among vascular risk factors, carotid atherosclerosis, and cortical volume and thickness in older adults.

BACKGROUND AND PURPOSE: The purpose of this study was to investigate whether the Framingham Cardiovascular Risk Profile and carotid artery intima-media thickness are associated with cortical volume and thickness. METHODS: Consecutive subjects participating in a prospective cohort study of aging and mild cognitive impairment enriched for vascular risk factors for atherosclerosis underwent structural MRI scans at 3-T and 4-T MRI at 3 sites. Freesurfer (Version 5.1) was used to obtain regional measures of neocortical volumes (mm3) and thickness (mm). Multiple linear regression was used to determine the association of Framingham Cardiovascular Risk Profile and carotid artery intima-media thickness with cortical volume and thickness. RESULTS: One hundred fifty-two subjects (82 men) were aged 78 (±7) years, 94 had a clinical dementia rating of 0, 58 had a clinical dementia rating of 0.5, and the mean Mini-Mental State Examination was 28±2. Framingham Cardiovascular Risk Profile score was inversely associated with total gray matter volume and parietal and temporal gray matter volume (adjusted P<0.04). Framingham Cardiovascular Risk Profile was inversely associated with parietal and total cerebral gray matter thickness (adjusted P<0.03). Carotid artery intima-media thickness was inversely associated with thickness of parietal gray matter only (adjusted P=0.04). Including history of myocardial infarction or stroke and radiological evidence of brain infarction, or apolipoprotein E genotype did not alter relationships with Framingham Cardiovascular Risk Profile or carotid artery intima-media thickness. CONCLUSIONS: Increased cardiovascular risk was associated with reduced gray matter volume and thickness in regions also affected by Alzheimer disease independent of infarcts and apolipoprotein E genotype. These results suggest a "double hit" toward developing dementia when someone with incipient Alzheimer disease also has high cardiovascular risk.

Atypical developmental trajectory of functionally significant cortical areas in children with chromosome 22q11.2 deletion syndrome.

Chromosome 22q11.2 deletion syndrome (22q11.2DS) is a neurogenetic disorder associated with neurocognitive impairments. This article focuses on the cortical gyrification changes that are associated with the genetic disorder in 6-15-year-old children with 22q11.2DS, when compared with a group of age-matched typically developing (TD) children. Local gyrification index (lGI; Schaer et al. [2008]: IEEE Trans Med Imaging 27:161-170) was used to characterize the cortical gyrification at each vertex of the pial surface. Vertex-wise statistical analysis of lGI differences between the two groups revealed cortical areas of significant reduction in cortical gyrification in children with 22q11.2DS, which were mainly distributed along the medial aspect of each hemisphere. To gain further insight into the developmental trajectory of the cortical gyrification, we examined age as a factor in lGI changes over the 6-15 years of development, within and across the two groups of children. Our primary results pertaining to the developmental trajectory of cortical gyrification revealed cortical regions where the change in lGI over the 6-15 years of age was significantly modulated by diagnosis, implying an atypical development of cortical gyrification in children with 22q11.2DS, when compared with the TD children. Significantly, these cortical areas included parietal structures that are associated, in typical individuals, with visuospatial, attentional, and numerical cognition tasks in which children with 22q11.2DS show impairments.

Volumetric correlates of spatiotemporal working and recognition memory impairment in aged rhesus monkeys.

Spatiotemporal and recognition memory are affected by aging in humans and macaque monkeys. To investigate whether these deficits are coupled with atrophy of memory-related brain regions, T(1)-weighted magnetic resonance images were acquired and volumes of the cerebrum, ventricles, prefrontal cortex (PFC), calcarine cortex, hippocampus, and striatum were quantified in young and aged rhesus monkeys. Subjects were tested on a spatiotemporal memory procedure (delayed response [DR]) that requires the integrity of the PFC and a medial temporal lobe-dependent recognition memory task (delayed nonmatching to sample [DNMS]). Region of interest analyses revealed that age inversely correlated with striatal, dorsolateral prefrontal cortex (dlPFC), and anterior cingulate cortex volumes. Hippocampal volume predicted acquisition of the DR task. Striatal volume correlated with DNMS acquisition, whereas total prefrontal gray matter, prefrontal white matter, and dlPFC volumes each predicted DNMS accuracy. A regional covariance analysis revealed that age-related volumetric changes could be captured in a distributed network that was coupled with declining performance across delays on the DNMS task. This volumetric analysis adds to growing evidence that cognitive aging in primates arises from region-specific morphometric alterations distributed across multiple memory-related brain systems, including subdivisions of the PFC.

Integrating conflict detection and attentional control mechanisms.

Human behavior involves monitoring and adjusting performance to meet established goals. Performance-monitoring systems that act by detecting conflict in stimulus and response processing have been hypothesized to influence cortical control systems to adjust and improve performance. Here we used fMRI to investigate the neural mechanisms of conflict monitoring and resolution during voluntary spatial attention. We tested the hypothesis that the ACC would be sensitive to conflict during attentional orienting and influence activity in the frontoparietal attentional control network that selectively modulates visual information processing. We found that activity in ACC increased monotonically with increasing attentional conflict. This increased conflict detection activity was correlated with both increased activity in the attentional control network and improved speed and accuracy from one trial to the next. These results establish a long hypothesized interaction between conflict detection systems and neural systems supporting voluntary control of visual attention.

Vigorous exercise increases brain lactate and Glx (glutamate+glutamine): a dynamic 1H-MRS study.

Vigorous exercise increases lactate and glucose uptake by the brain in excess of the increase in brain oxygen uptake. The metabolic fate of this non-oxidized carbohydrate entering the brain is poorly understood, but accumulation of lactate in the brain and/or increased net synthesis of amino acid neurotransmitters are possible explanations. Previous proton magnetic resonance spectroscopy (1H-MRS) studies using conventional pulse sequences have not detected changes in brain lactate following exercise. This contrasts with 1H-MRS studies showing increased brain lactate when blood lactate levels are raised by an intravenous infusion of sodium lactate. Using a J-editing 1H-MRS technique for measuring lactate, we demonstrated a significant 19% increase in lactate in the visual cortex following graded exercise to approximately 85% of predicted maximum heart rate. However, the magnitude of the increase was insufficient to account for more than a small fraction of the non-oxidized carbohydrate entering the brain with exercise. We also report a significant 18% increase in Glx (combined signal from glutamate and glutamine) in visual cortex following exercise, which may represent an activity-dependent increase in glutamate. Future studies will be necessary to test the hypothesis that non-oxidized carbohydrate entering the brain during vigorous exercise is directed, in part, toward increased net synthesis of amino acid neurotransmitters. The possible relevance of these findings to panic disorder and major depression is discussed.

Age-related regional network of magnetic resonance imaging gray matter in the rhesus macaque.

Human structural neuroimaging studies have supported the preferential effects of healthy aging on frontal cortex, but reductions in other brain regions have also been observed. We investigated the regional network pattern of gray matter using magnetic resonance imaging (MRI) in young adult and old rhesus macaques (RMs) to evaluate age effects throughout the brain in a nonhuman primate model of healthy aging in which the full complement of Alzheimer's disease (AD) pathology does not occur. Volumetric T1 MRI scans were spatially normalized and segmented for gray matter using statistical parametric mapping (SPM2) voxel-based morphometry. Multivariate network analysis using the scaled subprofile model identified a linear combination of two gray matter patterns that distinguished the young from old RMs. The combined pattern included reductions in bilateral dorsolateral and ventrolateral prefrontal and orbitofrontal and superior temporal sulcal regions with areas of relative preservation in vicinities of the cerebellum, globus pallidus, visual cortex, and parietal cortex in old compared with young RMs. Higher expression of this age-related gray matter pattern was associated with poorer performance in working memory. In the RM model of healthy aging, the major regionally distributed effects of advanced age on the brain involve reductions in prefrontal regions and in the vicinity of the superior temporal sulcus. The age-related differences in gray matter reflect the effects of healthy aging that cannot be attributed to AD pathology, providing support for the targeted effects of aging on the integrity of frontal lobe regions and selective temporal lobe areas and their associated cognitive functions.

Correlation of clinical and biochemical findings with diabetic ketoacidosis-related cerebral edema in children using magnetic resonance diffusion-weighted imaging.

OBJECTIVE: To determine clinical and biochemical factors influencing cerebral edema formation during diabetic ketoacidosis (DKA) in children. STUDY DESIGN: We used magnetic resonance diffusion-weighted imaging to quantify edema formation. We measured the apparent diffusion coefficient (ADC) of brain water during and after DKA treatment in 26 children and correlated ADC changes with clinical and biochemical variables. RESULTS: Mean ADC values were elevated during DKA treatment compared with baseline (8.13 +/- 0.47 vs 7.74 +/- 0.49 x 10(-4) mm(2)/sec, difference in means 0.40, 95% CI: 0.25 to 0.55, P < .001). Children with altered mental status during DKA had greater elevation in ADC. ADC elevation during DKA was positively correlated with initial serum urea nitrogen concentration (correlation coefficient 0.41, P = .03) and initial respiratory rate (correlation coefficient 0.61, P < .001). ADC elevation was not significantly correlated with initial serum glucose, sodium or effective osmolality, nor with changes in glucose, sodium or osmolality during treatment. Multivariable analyses identified the initial urea nitrogen concentration and respiratory rate as independently associated with ADC elevation. CONCLUSIONS: The degree of edema formation during DKA in children is correlated with the degree of dehydration and hyperventilation at presentation, but not with factors related to initial osmolality or osmotic changes during treatment. These data support the hypothesis that CE is related to cerebral hypoperfusion during DKA, and that osmotic fluctuations during DKA treatment do not play a primary causal role.

Attentional control and brain metabolite levels in methamphetamine abusers.

BACKGROUND: Methamphetamine abuse is associated with neurotoxicity to frontostriatal brain regions with concomitant deleterious effects on cognitive processes. METHODS: By using a computerized measure of selective attention and single-voxel proton magnetic resonance spectroscopy, we examined the relationship between attentional control and brain metabolite levels in the anterior cingulate cortex (ACC) and primary visual cortex (PVC) in 36 currently abstinent methamphetamine abusers and 16 non-substance-using controls. RESULTS: The methamphetamine abusers exhibited reduced attentional control (i.e., increased Stroop interference) compared with the controls (p = .04). Bonferroni-adjusted comparisons revealed that ACC levels of N-acetyl aspartate (NAA)-creatine and phosphocreatine (Cr) were lower and that levels of choline (Cho)-NAA were higher in the methamphetamine abusers compared with the controls, at the adjusted p value of .0125. Levels of NAA-Cr, but not of Cho-NAA, within the ACC correlated with measures of attentional control in the methamphetamine abusers (r = -.41; p = .01) but not in controls (r = .22; p = .42). No significant correlations were observed in the PVC (methamphetamine abusers, r = .19; p = .28, controls, r = .38; p = .15). CONCLUSIONS: Changes in neurochemicals within frontostriatal brain regions including ACC may contribute to deficits in attentional control among chronic methamphetamine abusers.

Methamphetamine users in sustained abstinence: a proton magnetic resonance spectroscopy study.

BACKGROUND: Abnormal patterns of metabolite levels have been detected by magnetic resonance spectroscopy in frontostriatal regions of individuals meeting DSM-IV criteria for methamphetamine dependence, but less is known about the effects of drug abstinence on metabolite levels. OBJECTIVE: To assess the effects of long-term methamphetamine use and drug abstinence on brain metabolite levels. DESIGN: To assess regional specific metabolite levels using magnetic resonance spectroscopy imaging techniques in 2 groups of currently abstinent methamphetamine users: methamphetamine users who recently initiated abstinence and methamphetamine users who had initiated abstinence more than 1 year prior to study. SETTING: Participants were recruited from outpatient substance abuse treatment centers. PARTICIPANTS: Eight methamphetamine users with sustained abstinence (1 year to 5 years) and 16 recently abstinent methamphetamine users (1 month to 6 months) were compared with 13 healthy, non-substance-using controls. MAIN OUTCOME MEASURES: Magnetic resonance spectroscopy measures of N-acetylaspartate-creatine and phosphocreatine (NAA/Cr), choline-creatine and phosphocreatine (Cho/Cr), and choline-N-acetylaspartate (Cho/NAA) ratios were obtained in the anterior cingulate cortex as well as in the primary visual cortex, which served as a control region. RESULTS: The absolute values of Cr did not differ between controls and methamphetamine users. Methamphetamine users had abnormally low NAA/Cr levels within the anterior cingulate cortex, regardless of the time spent abstinent (F(2,34) = 12.61; P<.001). No NAA/Cr group differences were observed in the primary visual cortex (F(2,33) = 0.29; P = .75). The Cho/NAA values for the anterior cingulate cortex were abnormally high in the methamphetamine users who recently initiated abstinence but followed a normal pattern in the methamphetamine users who had initiated abstinence more than 1 year prior to study (F(2,34) = 7.31; P = .002). CONCLUSIONS: The relative choline normalization across periods of abstinence suggests that following cessation of methamphetamine use, adaptive changes occur, which might contribute to some degree of normalization of neuronal structure and function in the anterior cingulum. More research is needed to elucidate the mechanisms underlying these adaptive changes.

Brain lactate responses during visual stimulation in fasting and hyperglycemic subjects: a proton magnetic resonance spectroscopy study at 1.5 Tesla.

Proton magnetic resonance spectroscopy ((1)H-MRS) studies showing increased lactate during neural activation support a broader role for lactate in brain energy metabolism than was traditionally recognized. Proton MRS measures of brain lactate responses have been used to study regional brain metabolism in clinical populations. This study examined whether variations in blood glucose influence the lactate response to visual stimulation in the visual cortex. Six subjects were scanned twice, receiving either saline or 21% glucose intravenously. Using (1)H-MRS at 1.5 Tesla with a long echo time (TE=288 ms), the lactate doublet was visible at 1.32 ppm in the visual cortex of all subjects. Lactate increased significantly from resting to visual stimulation. Hyperglycemia had no effect on this increase. The order of the slice-selective gradients for defining the spectroscopy voxel had a pronounced effect on the extent of contamination by signal originating outside the voxel. The results of this preliminary study demonstrate a method for observing a consistent activity-stimulated increase in brain lactate at 1.5 T and show that variations in blood glucose across the normal range have little effect on this response.

Longitudinal analysis of the developing rhesus monkey brain using magnetic resonance imaging: birth to adulthood.

We have longitudinally assessed normative brain growth patterns in naturalistically reared Macaca mulatta monkeys. Postnatal to early adulthood brain development in two cohorts of rhesus monkeys was analyzed using magnetic resonance imaging. Cohort A consisted of 24 rhesus monkeys (12 male, 12 female) and cohort B of 21 monkeys (11 male, 10 female). All subjects were scanned at 1, 4, 8, 13, 26, 39, and 52 weeks; cohort A had additional scans at 156 weeks (3 years) and 260 weeks (5 years). Age-specific segmentation templates were developed for automated volumetric analyses of the T1-weighted magnetic resonance imaging scans. Trajectories of total brain size as well as cerebral and subcortical subdivisions were evaluated over this period. Total brain volume was about 64 % of adult estimates in the 1-week-old monkey. Brain volume of the male subjects was always, on average, larger than the female subjects. While brain volume generally increased between any two imaging time points, there was a transient plateau of brain growth between 26 and 39 weeks in both cohorts of monkeys. The trajectory of enlargement differed across cortical regions with the occipital cortex demonstrating the most idiosyncratic pattern of maturation and the frontal and temporal lobes showing the greatest and most protracted growth. A variety of allometric measurements were also acquired and body weight gain was most closely associated with the rate of brain growth. These findings provide a valuable baseline for the effects of fetal and early postnatal manipulations on the pattern of abnormal brain growth related to neurodevelopmental disorders.

The amygdala is enlarged in children but not adolescents with autism; the hippocampus is enlarged at all ages.

Autism is a neurodevelopmental disorder characterized by impairments in reciprocal social interaction, deficits in verbal and nonverbal communication, and a restricted repertoire of activities or interests. We performed a magnetic resonance imaging study to better define the neuropathology of autistic spectrum disorders. Here we report findings on the amygdala and the hippocampal formation. Borders of the amygdala, hippocampus, and cerebrum were defined, and their volumes were measured in male children (7.5-18.5 years of age) in four diagnostic groups: autism with mental retardation, autism without mental retardation, Asperger syndrome, and age-matched typically developing controls. Although there were no differences between groups in terms of total cerebral volume, children with autism (7.5-12.5 years of age) had larger right and left amygdala volumes than control children. There were no differences in amygdala volume between the adolescent groups (12.75-18.5 years of age). Interestingly, the amygdala in typically developing children increases substantially in volume from 7.5 to 18.5 years of age. Thus, the amygdala in children with autism is initially larger, but does not undergo the age-related increase observed in typically developing children. Children with autism, with and without mental retardation, also had a larger right hippocampal volume than typically developing controls, even after controlling for total cerebral volume. Children with autism but without mental retardation also had a larger left hippocampal volume relative to controls. These cross-sectional findings indicate an abnormal program of early amygdala development in autism and an abnormal pattern of hippocampal development that persists through adolescence. The cause of amygdala and hippocampal abnormalities in autism is currently unknown.

Investigation of neuroanatomical differences between autism and Asperger syndrome.

BACKGROUND: Autism and Asperger syndrome (ASP) are neurobiological conditions with overlapping behavioral symptoms and of unknown etiologies. Results from previous autism neuroimaging studies have been difficult to replicate, possibly owing to site differences in subject samples, scanning procedures, and image-processing methods. We sought (1) to determine whether low-functioning autism (LFA; IQ<70), high-functioning autism (HFA; IQ>or=70), and ASP constitute distinct biological entities as evidenced by neuroanatomical measures, and (2) to assess for intersite differences. METHODS: Case-control study examining coronally oriented 124-section spoiled gradient echo images acquired on 3 magnetic resonance imaging (MRI) systems, and processed by BrainImage 5.X. Participants were recruited and underwent scanning at 2 academic medicine departments. Participants included 4 age-matched groups of volunteer boys aged 7.8 to 17.9 years (13 patients with LFA, 18 with HFA, 21 with ASP, and 21 control subjects), and 3 volunteer adults for neuroimaging reliability. Main outcome measures included volumetric measures of total, white, and gray matter for cerebral and cerebellar tissues. RESULTS: Intersite differences were seen for subject age, IQ, and cerebellum measures. Cerebral gray matter volume was enlarged in both HFA and LFA compared with controls (P =.009 and P =.04, respectively). Cerebral gray matter volume in ASP was intermediate between that of HFA and controls, but nonsignificant. Exploratory analyses revealed a negative correlation between cerebral gray matter volume and performance IQ within HFA but not ASP. A positive correlation between cerebral white matter volume and performance IQ was observed within ASP but not HFA. CONCLUSIONS: Lack of replication between previous autism MRI studies could be due to intersite differences in MRI systems and subjects' age and IQ. Cerebral gray tissue findings suggest that ASP is on the mild end of the autism spectrum. However, exploratory assessments of brain-IQ relationships reveal differences between HFA and ASP, indicating that these conditions may be neurodevelopmentally different when patterns of multiple measures are examined. Further investigations of brain-behavior relationships are indicated to confirm these findings.

Detection of cerebral {beta}-hydroxy butyrate, acetoacetate, and lactate on proton MR spectroscopy in children with diabetic ketoacidosis.

BACKGROUND AND PURPOSE: Ketone bodies provide important alternate fuel for brain metabolism, and their transport into the brain increases with prolonged fasting. During diabetic ketoacidosis (DKA), serum ketone concentrations markedly increase; however, little is known about whether ketone bodies accumulate in cerebral tissues during DKA. We used proton MR spectroscopy (MRS) to detect cerebral beta-hydroxy butyrate (betaOHB) and acetone/acetocaetate (AcAc) in children with DKA. METHODS: Twenty-five children underwent brain MRS: nine within 4 hours of the start of treatment for DKA; 11, at 4-8 hours; and five, at 8-12 hours. MRS was repeated after their recovery from the DKA episode at > or =72 hours after the start of treatment. MRS was evaluated for peaks corresponding to betaOHB (doublet centered on 1.20 ppm) and lactate (doublet centered on 1.33 ppm). Difference spectroscopy was used to identify the AcAc peak at 2.22-2.26 ppm. RESULTS: betaOHB was detected in 13 children (52%), more frequently within 4 hours (eight children, 89%) than after 4 hours (five children, 31%). AcAc was detected in 15 children (60%), more frequently at >4 hours after the start of treatment (12 patients, 75%) than in the first 4 hours (three patients, 33%). Lactate was detected in five children (18%), all within the first 8 hours of treatment. CONCLUSION: In children, betaOHB and AcAc accumulate in the brain during DKA, and they can be detected on MRS. Care should be taken in interpreting MRS results in patients with DKA to avoid erroneously attributing betaOHB peaks to lactate.

Estimation of extraction fraction (EF) and glomerular filtration rate (GFR) using MRI: considerations derived from a new Gd-chelate biodistribution model simulation.

Previous reports have described the use of magnetic resonance imaging (MRI) to estimate single-kidney extraction fraction (EF) and glomerular filtration rate (GFR), by measuring the concentration difference of intravenously injected Gd-chelate ([Gd]) in the renal artery and renal vein from measurements of blood T1. Problematic is the fact that [Gd] measurements in the renal artery are often inaccurate due to the small size, tortuousness and motion of the vessel. Consequently, the [Gd] in the inferior vena cava (IVC) below the renal vein ostia (i.e., the infrarenal IVC) has been used instead of the renal artery [Gd], based on the assumption that the [Gd] in the infrarenal IVC is the same as it is in the renal artery. However, this assumption has neither been theoretically nor experimentally investigated. Herein, we describe new difference and differential equation pharmacological models that can predict the biodistribution of Gd-chelate throughout the extracellular space. Assuming known average normal blood flows and GFR, our models predict that the infrarenal IVC [Gd] is 3.2% to 4.7% greater than the renal artery [Gd], and that the EF estimate using this IVC measurement is overestimated by 14.2%-20.0%. To support these predictions, algebraic equations are derived which show that the infrarenal IVC must develop a relatively high [Gd] in order to satisfy Gd flux constraints within the vascular system. These results suggest that the infrarenal IVC [Gd] is not a valid substitute for the renal artery [Gd].

Magnetic resonance spectroscopy of the brain: a review of physical principles and technical methods.

Magnetic resonance spectroscopy (MRS) provides unique information about the neurobiological substrates of brain function in health and disease. However, many of the physical principles underlying MRS are distinct from those underlying magnetic resonance imaging, and they may not be widely understood by neuroscientists new to this methodology. This review describes these physical principles and many of the technical methods in current use for MRS experiments. A better understanding these principles and methods may help investigators select pulse sequences and quantification methods best suited to the aims of their research program and avoid pitfalls that can hamper new investigators in this field.