Robust hyperpolarized 13C metabolic imaging with selective non‐excitation of pyruvate (SNEP)

In vivo metabolic imaging using hyperpolarized [1‐¹³C]pyruvate provides localized biochemical information and is particularly useful in detecting early disease changes, as well as monitoring disease progression and treatment response. However, a major limitation of hyperpolarized magnetization is its unrecoverable decay, due not only to T₁ relaxation but also to radio‐frequency (RF) excitation. RF excitation schemes used in metabolic imaging must therefore be able to utilize available hyperpolarized magnetization efficiently and robustly for the optimal detection of substrate and metabolite activities. In this work, a novel RF excitation scheme called selective non‐excitation of pyruvate (SNEP) is presented. This excitation scheme involves the use of a spectral selective RF pulse to specifically exclude the excitation of [1‐¹³C]pyruvate, while uniformly exciting the key metabolites of interest (namely [1‐¹³C]lactate and [1‐¹³C]alanine) and [1‐¹³C]pyruvate‐hydrate. By eliminating the loss of hyperpolarized [1‐¹³C]pyruvate magnetization due to RF excitation, the signal from downstream metabolite pools is increased together with enhanced dynamic range. Simulation results, together with phantom measurements and in vivo experiments, demonstrated the improvement in signal‐to‐noise ratio (SNR) and the extension of the lifetime of the [1‐¹³C]lactate and [1‐¹³C]alanine pools when compared with conventional non‐spectral selective (NS) excitation. SNEP has also been shown to perform comparably well with multi‐band (MB) excitation, yet SNEP possesses distinct advantages, including ease of implementation, less stringent demands on gradient performance, increased robustness to frequency drifts and B₀ inhomogeneity as well as easier quantification involving the use of [1‐¹³C]pyruvate‐hydrate as a proxy for the actual [1‐¹³C] pyruvate signal. SNEP is therefore a promising alternative for robust hyperpolarized [1‐¹³C]pyruvate metabolic imaging with high fidelity. Copyright © 2015 John Wiley & Sons, Ltd.     

A referenceless Nyquist ghost correction workflow for echo planar imaging of hyperpolarized [1‐13C]pyruvate and [1‐13C]lactate

Single‐shot echo planar imaging (EPI), which allows an image to be acquired using a single excitation pulse, is used widely for imaging the metabolism of hyperpolarized ¹³C‐labelled metabolites in vivo as the technique is rapid and minimizes the depletion of the hyperpolarized signal. However, EPI suffers from Nyquist ghosting, which normally is corrected for by acquiring a reference scan. In a dynamic acquisition of a series of images, this results in the sacrifice of a time point if the reference scan involves a full readout train with no phase encoding. This time penalty is negligible if an integrated navigator echo is used, but at the cost of a lower signal‐to‐noise ratio (SNR) as a result of prolonged T₂* decay. We describe here a workflow for hyperpolarized ¹³C EPI that requires no reference scan. This involves the selection of a ghost‐containing background from a ¹³C image of a single metabolite at a single time point, the identification of phase correction coefficients that minimize signal in the selected area, and the application of these coefficients to images acquired at all time points and from all metabolites. The workflow was compared in phantom experiments with phase correction using a ¹³C reference scan, and yielded similar results in situations with a regular field of view (FOV), a restricted FOV and where there were multiple signal sources. When compared with alternative phase correction methods, the workflow showed an SNR benefit relative to integrated ¹³C reference echoes (>15%) or better ghost removal relative to a ¹H reference scan. The residual ghosting in a slightly de‐shimmed B₀ field was 1.6% using the proposed workflow and 3.8% using a ¹H reference scan. The workflow was implemented with a series of dynamically acquired hyperpolarized [1‐¹³C]pyruvate and [1‐¹³C]lactate images in vivo, resulting in images with no observable ghosting and which were quantitatively similar to images corrected using a ¹³C reference scan.     

Apparent rate constant mapping using hyperpolarized [1–13C]pyruvate

Hyperpolarization of [1‐13C]pyruvate in solution allows real‐time measurement of uptake and metabolism using MR spectroscopic methods. After injection and perfusion, pyruvate is taken up by the cells and enzymatically metabolized into downstream metabolites such as lactate, alanine, and bicarbonate. In this work, we present comprehensive methods for the quantification and interpretation of hyperpolarized 13C metabolite signals. First, a time‐domain spectral fitting method is described for the decomposition of FID signals into their metabolic constituents. For this purpose, the required chemical shift frequencies are automatically estimated using a matching pursuit algorithm. Second, a time‐discretized formulation of the two‐site exchange kinetic model is used to quantify metabolite signal dynamics by two characteristic rate constants in the form of (i) an apparent build‐up rate (quantifying the build‐up of downstream metabolites from the pyruvate substrate) and (ii) an effective decay rate (summarizing signal depletion due to repetitive excitation, T1‐relaxation and backward conversion). The presented spectral and kinetic quantification were experimentally verified in vitro and in vivo using hyperpolarized [1‐13C]pyruvate. Using temporally resolved IDEAL spiral CSI, spatially resolved apparent rate constant maps are also extracted. In comparison to single metabolite images, apparent build‐up rate constant maps provide improved contrast by emphasizing metabolically active tissues (e.g. tumors) and suppression of high perfusion regions with low conversion (e.g. blood vessels). Apparent build‐up rate constant mapping provides a novel quantitative image contrast for the characterization of metabolic activity. Its possible implementation as a quantitative standard will be subject to further studies. Copyright © 2014 John Wiley & Sons, Ltd.     

An indirect method for in vivo T2 mapping of [1‐13C] pyruvate using hyperpolarized 13C CSI

An indirect method for in vivo T₂ mapping of ¹³C–labeled metabolites using T₂ and T₂* information of water protons obtained a priori is proposed. The T₂ values of ¹³C metabolites are inferred using the relationship to T₂′ of coexisting ¹H and the T₂* of ¹³C metabolites, which is measured using routine hyperpolarized ¹³C CSI data. The concept is verified with phantom studies. Simulations were performed to evaluate the extent of T₂ estimation accuracy due to errors in the other measurements. Also, bias in the ¹³C T₂* estimation from the ¹³C CSI data was studied. In vivo experiments were performed from the brains of normal rats and a rat with C6 glioma. Simulation results indicate that the proposed method provides accurate and unbiased ¹³C T₂ values within typical experimental settings. The in vivo studies found that the estimated T₂ of [1‐¹³C] pyruvate using the indirect method was longer in tumor than in normal tissues and gave values similar to previous reports. This method can estimate localized T₂ relaxation times from multiple voxels using conventional hyperpolarized ¹³C CSI and can potentially be used with time resolved fast CSI.     

Assessing the influence of isoflurane anesthesia on cardiac metabolism using hyperpolarized [1‐13C]pyruvate

Isoflurane is a frequently used anesthetic in small‐animal dissolution dynamic nuclear polarization‐magnetic resonance imaging (DNP‐MRI) studies. Although the literature suggests interactions with mitochondrial metabolism, the influence of the compound on cardiac metabolism has not been assessed systematically to date. In the present study, the impact of low versus high isoflurane concentration was examined in a crossover experiment in healthy rats. The results revealed that cardiac metabolism is modulated by isoflurane concentration, showing increased [1‐¹³C]lactate and reduced [¹³C]bicarbonate production during high isoflurane relative to low isoflurane dose [average differences: +16% [1‐¹³C]lactate/total myocardial carbon, –22% [¹³C]bicarbonate/total myocardial carbon; +51% [1‐¹³C]lactate/[¹³C]bicarbonate]. These findings emphasize that reproducible anesthesia is important when studying cardiac metabolism. As the depth of anesthesia is difficult to control in an experimental animal setting, careful study design is required to exclude confounding factors.     

Voxel‐by‐voxel correlations of perfusion,substrate, and metabolite signals in dynamic hyperpolarized 13C imaging

In this study, a mixture of pyruvic acid and the perfusion agent HP001 was co‐polarized for simultaneous assessment of perfusion and metabolism in vivo. The pre‐polarized mixture was administered to rats with subcutaneous MDA‐MB‐231 breast cancer xenografts and imaged using an interleaved sequence with designed spectral–spatial pulses and flyback echo‐planar readouts. Voxel‐by‐voxel signal correlations from 10 animals (15 data sets) were analyzed for tumour, kidney, and muscle regions of interest. The relationship between perfusion and hyperpolarized signal was explored on a voxel‐by‐voxel basis in various metabolically active tissues, including tumour, healthy kidneys, and skeletal muscle. Positive pairwise correlations between lactate, pyruvate, and HP001 observed in all 10 tumours suggested that substrate delivery was the dominant factor limiting the conversion of pyruvate to lactate in the tumour model used in this study. On the other hand, in cases where conversion is the limiting factor, such as in healthy kidneys, both pyruvate and lactate can act as excellent perfusion markers. In intermediate cases between the two limits, such as in skeletal muscle, some perfusion information may be inferred from the (pyruvate + lactate) signal distribution. Co‐administration of pyruvate with a dynamic nuclear polarization (DNP) perfusion agent is an effective approach for distinguishing between slow metabolism and poor perfusion and a practical strategy for lactate signal normalization to account for substrate delivery, especially in cases of rapid pyruvate‐to‐lactate conversion and in poorly perfused regions with inadequate pyruvate signal‐to‐noise ratio for reliable determination of the lactate‐to‐pyruvate ratio. Copyright © 2016 John Wiley & Sons, Ltd.     

Comparison of acquisition schemes for hyperpolarised 13C imaging

The aim of this study was to characterise and compare widely used acquisition strategies for hyperpolarised ¹³C imaging. Free induction decay chemical shift imaging (FIDCSI), echo‐planar spectroscopic imaging (EPSI), IDEAL spiral chemical shift imaging (ISPCSI) and spiral chemical shift imaging (SPCSI) sequences were designed for two different regimes of spatial resolution. Their characteristics were studied in simulations and in tumour‐bearing rats after injection of hyperpolarised [1‐¹³C]pyruvate on a clinical 3‐T scanner. Two or three different sequences were used on the same rat in random order for direct comparison. The experimentally obtained lactate signal‐to‐noise ratio (SNR) in the tumour matched the simulations. Differences between the sequences were mainly found in the encoding efficiency, gradient demand and artefact behaviour. Although ISPCSI and SPCSI offer high encoding efficiencies, these non‐Cartesian trajectories are more prone than EPSI and FIDCSI to artefacts from various sources. If the encoding efficiency is sufficient for the desired application, EPSI has been proven to be a robust choice. Otherwise, faster spiral acquisition schemes are recommended. The conclusions found in this work can be applied directly to clinical applications. Copyright © 2015 John Wiley & Sons, Ltd.     

1H NMR and hyperpolarized 13C NMR assays of pyruvate–lactate: a comparative study

Pyruvate–lactate exchange is mediated by the enzyme lactate dehydrogenase (LDH) and is central to the altered energy metabolism in cancer cells. The measurement of exchange kinetics using hyperpolarized ¹³C NMR has provided a biomarker of response to novel therapeutics. However, the observable signal is restricted to the exchanging hyperpolarized ¹³C pools and the endogenous pools of ¹²C‐labelled metabolites are invisible in these measurements. In this study, we investigated an alternative in vitro ¹H NMR assay, using [3‐¹³C]pyruvate, and compared the measured kinetics with a hyperpolarized ¹³C NMR assay, using [1‐¹³C]pyruvate, under the same conditions in human colorectal carcinoma SW1222 cells. The apparent forward reaction rate constants (k_PL) derived from the two assays showed no significant difference, and both assays had similar reproducibility (k_PL = 0.506 ± 0.054 and k_PL = 0.441 ± 0.090 nmol/s/10⁶ cells; mean ± standard deviation; n = 3); ¹H, ¹³C assays, respectively). The apparent backward reaction rate constant (k_LP) could only be measured with good reproducibility using the ¹H NMR assay (k_LP = 0.376 ± 0.091 nmol/s/10⁶ cells; mean ± standard deviation; n = 3). The ¹H NMR assay has adequate sensitivity to measure real‐time pyruvate–lactate exchange kinetics in vitro, offering a complementary and accessible assay of apparent LDH activity. Copyright © 2013 John Wiley & Sons, Ltd.     

Pulse sequence considerations for quantification of pyruvate‐to‐lactate conversion kPL in hyperpolarized 13C imaging

Hyperpolarized ¹³C MRI takes advantage of the unprecedented 50 000‐fold signal‐to‐noise ratio enhancement to interrogate cancer metabolism in patients and animals. It can measure the pyruvate‐to‐lactate conversion rate, k_PL, a metabolic biomarker of cancer aggressiveness and progression. Therefore, it is crucial to evaluate k_PL reliably. In this study, three sequence components and parameters that modulate k_PL estimation were identified and investigated in model simulations and through in vivo animal studies using several specifically designed pulse sequences. These factors included a magnetization spoiling effect due to RF pulses, a crusher gradient‐induced flow suppression, and intrinsic image weightings due to relaxation. Simulation showed that the RF‐induced magnetization spoiling can be substantially improved using an inputless k_PL fitting. In vivo studies found a significantly higher apparent k_PL with an additional gradient that leads to flow suppression (k_{PL,FID‐Delay,Crush}/k_{PL,FID‐Delay} = 1.37 ± 0.33, P < 0.01, N = 6), which agrees with simulation outcomes (12.5% k_PL error with Δv = 40 cm/s), indicating that the gradients predominantly suppressed flowing pyruvate spins. Significantly lower k_PL was found using a delayed free induction decay (FID) acquisition versus a minimum‐T_E version (k_{PL,FID‐Delay}/k_PL,FID = 0.67 ± 0.09, P < 0.01, N = 5), and the lactate peak had broader linewidth than pyruvate (Δω_lactate/Δω_pyruvate = 1.32 ± 0.07, P < 0.000 01, N = 13). This illustrated that lactate's T₂*, shorter than that of pyruvate, can affect calculated k_PL values. We also found that an FID sequence yielded significantly lower k_PL versus a double spin‐echo sequence that includes spin‐echo spoiling, flow suppression from crusher gradients, and more T₂ weighting (k_PL,DSE/k_PL,FID = 2.40 ± 0.98, P < 0.0001, N = 7). In summary, the pulse sequence, as well as its interaction with pharmacokinetics and the tissue microenvironment, can impact and be optimized for the measurement of k_PL. The data acquisition and analysis pipelines can work synergistically to provide more robust and reproducible k_PL measures for future preclinical and clinical studies.     

Detection of lung transplant rejection in a rat model using hyperpolarized [1‐13C] pyruvate‐based metabolic imaging

The current standard for noninvasive imaging of acute rejection consists of X‐ray/CT, which derive their contrast from changes in ventilation, inflammation and edema, as well as remodeling during rejection. We propose the use of hyperpolarized [1‐¹³C] pyruvate MRI—which provides real‐time metabolic assessment of tissue—as an early biomarker for tissue rejection. In this preliminary study, we used μCT‐derived parameters and HP ¹³C MR‐derived biomarkers to predict rejection in an orthotopic left lung transplant model in both allogeneic and syngeneic rats. On day 3, the normalized lung density—a parameter that accounts for both lung volume (mL) and density (HU)—was ?0.335 (CI: ‐0.598, ?0.073) and ? 0.473 (CI: ‐0.726, ?0.220) for the allograft and isograft, respectively (not significant, 0.40). The lactate‐to‐pyruvate ratios—derived from the HP ¹³C MRI—for the allograft and isograft were 0.200 (CI: 0.161, 0.240) and 0.114 (CI: 0.074, 0.153), respectively (significant, 0.020). Both techniques showed tissue rejection on day 7. A separate sub‐study revealed CD8+ cells as the primary source of the lactate‐to‐pyruvate signal. Our study suggests that hyperpolarized (HP) [1‐¹³C] pyruvate MRI is a promising early biomarker for tissue rejection that provides metabolic assessment in real time based on changes in cellularity and metabolism of lung tissue and the infiltrating inflammatory cells, and may be able to predict tissue rejection earlier than X‐ray/CT.     

Metabolic imaging of acute and chronic infarction in the perfused rat heart using hyperpolarised [1‐13C]pyruvate

Hyperpolarised ¹³C MRI can be used to generate metabolic images of the heart in vivo. However, there have been no similar studies performed in the isolated perfused heart. Therefore, the aim of this study was to develop a method for the creation of ¹³C metabolite maps of the perfused rat heart and to demonstrate the technique in a study of acute and chronic myocardial infarction. Male Wistar rat hearts were isolated, perfused and imaged before and after occlusion of the left anterior descending (LAD) coronary artery, creating an acute infarct group. In addition, a chronic infarct group was generated from hearts which had their LAD coronary artery occluded in vivo. Four weeks later, hearts were excised, perfused and imaged to generate metabolic maps of infused pyruvate and its metabolites lactate and bicarbonate. Myocardial perfusion and energetics were assessed by first‐pass perfusion imaging and ³¹P MRS, respectively. In both acute and chronically infarcted hearts, perfusion was reduced to the infarct region, as revealed by reduced gadolinium influx and lower signal intensity in the hyperpolarised pyruvate images. In the acute infarct region, there were significant alterations in the lactate (increased) and bicarbonate (decreased) signal ratios. In the chronically infarcted region, there was a significant reduction in both bicarbonate and lactate signals. ³¹P‐derived energetics revealed a significant decrease between control and chronic infarcted hearts. Significant decreases in contractile function between control and both acute and chronic infracted hearts were also seen. In conclusion, we have demonstrated that hyperpolarised pyruvate can detect reduced perfusion in the rat heart following both acute and chronic infarction. Changes in lactate and bicarbonate ratios indicate increased anaerobic metabolism in the acute infarct, which is not observed in the chronic infarct. Thus, this study has successfully demonstrated a novel imaging approach to assess altered metabolism in the isolated perfused rat heart. © 2013 The Authors. NMR in Biomedicine published by John Wiley & Sons Ltd     

Simultaneous imaging of 13C metabolism and 1H structure: technical considerations and potential applications

Real‐time imaging of ¹³C metabolism in vivo has been enabled by recent advances in hyperpolarization. As a result of the inherently low natural abundance of endogenous ¹³C nuclei, hyperpolarized ¹³C images lack structural information that could be used to aid in motion detection and anatomical registration. Motion before or during the ¹³C acquisition can therefore result in artifacts and misregistration that may obscure measures of metabolism. In this work, we demonstrate a method to simultaneously image both ¹H and ¹³C nuclei using a dual‐nucleus spectral–spatial radiofrequency excitation and a fully coincident readout for rapid multinuclear spectroscopic imaging. With the appropriate multinuclear hardware, and the means to simultaneously excite and receive on both channels, this technique is straightforward to implement requiring little to no increase in scan time. Phantom and in vivo experiments were performed with both Cartesian and spiral trajectories to validate and illustrate the utility of simultaneous acquisitions. Motion compensation of dynamic metabolic measurements acquired during free breathing was demonstrated using motion tracking derived from ¹H data. Simultaneous multinuclear imaging provides structural ¹H and metabolic ¹³C images that are correlated both spatially and temporally, and are therefore amenable to joint ¹H and ¹³C analysis and correction of structure–function images. Copyright © 2015 John Wiley & Sons, Ltd.     

Real‐time ALT and LDH activities determined in viable precision‐cut mouse liver slices using hyperpolarized [1‐13C]pyruvate—Implications for studies on biopsied liver tissues

Precision‐cut liver slices (PCLS) are widely used in liver research as they provide a liver model with all liver cell types in their natural architecture. The purpose of this study was to demonstrate the use of PCLS for hyperpolarized metabolic investigation in a mouse model, for potential future application in liver biopsy cores. Fresh normal liver was harvested from six mice. 500 μm PCLS were prepared and placed in a 10 mm NMR tube in an NMR spectrometer and perfused continuously. ³¹P spectra were acquired to evaluate the presence of adenosine triphosphate (ATP) and validate viability in all samples. Hyperpolarized [1‐¹³C]pyruvate was flushed into the NMR tube in the spectrometer. Consecutive ¹³C NMR spectra were acquired immediately after the injection using both non‐selective (five injections, two livers) and selective RF excitation (six injections, three livers). The ³¹P spectra showed the characteristic signals of ATP, confirming the viability of the PCLS for more than 2.5 h in the spectrometer. After each of the [1‐¹³C]pyruvate injections, both [1‐¹³C]lactate and [1‐¹³C]alanine signals were detected. Selective RF excitation aimed at both [1‐¹³C]lactate and [1‐¹³C]alanine enabled better visualization and quantification of the metabolic activity. Using this acquisition approach only the newly formed metabolites are observed upon excitation, and their intensities relative to those of hyperpolarized pyruvate enable quantification of metabolite production rates. This rate of lactate and alanine production appeared to be constant throughout the measurement time, with alanine production about 2.3 times higher than lactate. In summary, the viability of PCLS in an NMR spectrometer was demonstrated and hyperpolarized [1‐¹³C]pyruvate metabolism was recorded. This study opens up the possibility of evaluating alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) activities in human liver biopsies, while preserving the tissue architecture and viability. In healthy, well‐perfused liver slices the ratio of ALT to LDH activity is about 2.3.     

Simultaneous investigation of cardiac pyruvate dehydrogenase flux, Krebs cycle metabolism and pH, using hyperpolarized [1,2-(13)C2]pyruvate in vivo

(13)C MR spectroscopy studies performed on hearts ex vivo and in vivo following perfusion of prepolarized [1-(13)C]pyruvate have shown that changes in pyruvate dehydrogenase (PDH) flux may be monitored non-invasively. However, to allow investigation of Krebs cycle metabolism, the (13)C label must be placed on the C2 position of pyruvate. Thus, the utilization of either C1 or C2 labeled prepolarized pyruvate as a tracer can only afford a partial view of cardiac pyruvate metabolism in health and disease. If the prepolarized pyruvate molecules were labeled at both C1 and C2 positions, then it would be possible to observe the downstream metabolites that were the results of both PDH flux ((13)CO(2) and H(13)CO(3)(-)) and Krebs cycle flux ([5-(13)C]glutamate) with a single dose of the agent. Cardiac pH could also be monitored in the same experiment, but adequate SNR of the (13)CO(2) resonance may be difficult to obtain in vivo. Using an interleaved selective RF pulse acquisition scheme to improve (13)CO(2) detection, the feasibility of using dual-labeled hyperpolarized [1,2-(13)C(2)]pyruvate as a substrate for dynamic cardiac metabolic MRS studies to allow simultaneous investigation of PDH flux, Krebs cycle flux and pH, was demonstrated in vivo.     

Simultaneous imaging of hyperpolarized [1,4‐13C2]fumarate, [1‐13C]pyruvate and 18F–FDG in a rat model of necrosis in a clinical PET/MR scanner

A co‐polarization scheme for [1,4‐¹³C₂]fumarate and [1‐¹³C]pyruvate is presented to simultaneously assess necrosis and metabolism in rats with hyperpolarized ¹³C magnetic resonance (MR). The co‐polarization was performed in a SPINlab polarizer. In addition, the feasibility of simultaneous positron emission tomography (PET) and MR of small animals with a clinical PET/MR scanner is demonstrated. The hyperpolarized metabolic MR and PET was demonstrated in a rat model of necrosis. The polarization and T₁ of the co‐polarized [1,4‐¹³C₂]fumarate and [1‐¹³C]pyruvate substrates were measured in vitro and compared with those obtained when the substrates were polarized individually. A polarization of 36 ± 4% for fumarate and 37 ± 6% for pyruvate was obtained. We found no significant difference in the polarization and T₁ values between the dual and single substrate polarization. Rats weighing about 400 g were injected intramuscularly in one of the hind legs with 200 μL of turpentine to induce necrosis. Two hours later, ¹³C metabolic maps were obtained with a chemical shift imaging sequence (16 × 16) with a resolution of 3.1 × 5.0 × 25.0 mm³. The ¹³C spectroscopic images were acquired in 12 s, followed by an 8‐min ¹⁸F‐2‐fluoro‐2‐deoxy‐d ‐glucose (¹⁸F–FDG) PET acquisition with a resolution of 3.5 mm. [1,4‐¹³C₂]Malate was observed from the tissue injected with turpentine indicating necrosis. Normal [1‐¹³C]pyruvate metabolism and ¹⁸F–FDG uptake were observed from the same tissue. The proposed co‐polarization scheme provides a means to utilize multiple imaging agents simultaneously, and thus to probe various metabolic pathways in a single examination. Moreover, it demonstrates the feasibility of small animal research on a clinical PET/MR scanner for combined PET and hyperpolarized metabolic MR.     

Cortex‐sparing fiber dissection: an improved method for the study of white matter anatomy in the human brain

Classical fiber dissection of post mortem human brains enables us to isolate a fiber tract by removing the cortex and overlying white matter. In the current work, a modification of the dissection methodology is presented that preserves the cortex and the relationships within the brain during all stages of dissection, i.e. ‘cortex‐sparing fiber dissection’. Thirty post mortem human hemispheres (15 right side and 15 left side) were dissected using cortex‐sparing fiber dissection. Magnetic resonance imaging study of a healthy brain was analyzed using diffusion tensor imaging (DTI)‐based tractography software. DTI fiber tract reconstructions were compared with cortex‐sparing fiber dissection results. The fibers of the superior longitudinal fasciculus (SLF), inferior fronto‐occipital fasciculus (IFOF), inferior longitudinal fasciculus (ILF) and uncinate fasciculus (UF) were isolated so as to enable identification of their cortical terminations. Two segments of the SLF were identified: first, an indirect and superficial component composed of a horizontal and vertical segment; and second, a direct and deep component or arcuate fasciculus. The IFOF runs within the insula, temporal stem and sagittal stratum, and connects the frontal operculum with the occipital, parietal and temporo‐basal cortex. The UF crosses the limen insulae and connects the orbito‐frontal gyri with the anterior temporal lobe. Finally, a portion of the ILF was isolated connecting the fusiform gyrus with the occipital gyri. These results indicate that cortex‐sparing fiber dissection facilitates study of the 3D anatomy of human brain tracts, enabling the tracing of fibers to their terminations in the cortex. Consequently, it is an important tool for neurosurgical training and neuroanatomical research.