Mapping plasticity in the forepaw digit barrel subfield of rat brains using functional MRI.

The topographic organization of the forepaw barrel subfield in layer IV of rat primary somatosensory cortex (S1) is a good model for studying neural function and plasticity. The goal of this study was to test the feasibility of functional MRI (fMRI) to map the forepaw digit representations in the S1 of the rat and its plasticity after digit amputation. Three dimensional echo-planar imaging with 300 micron isotropic resolution at 11.7 T was used to achieve high signal-to-noise ratios and laminar layer resolution. By alternating electrical stimulation of the 2nd (D2) and 4th (D4) digits, functional activation in layer IV of the barrel subfields could be distinguished using a differential analysis. Furthermore, 2 and a half months after the amputation of the 3rd digit in baby rats, the overlapping area between D2 and D4 representations was increased. This indicates that the forepaw barrel subfield previously associated with the ablated digit is now associated with the representation of nearby digits, which is consistent with studies using electrophysiology and cytochrome oxidase staining.

Mapping cortical representations of the rodent forepaw and hindpaw with BOLD fMRI reveals two spatial boundaries.

Electrical stimulation of the rat forepaw and hindpaw was employed to study the spatial distribution of BOLD fMRI. Averaging of multiple fMRI sessions significantly improved the spatial stability of the BOLD signal and enabled quantitative determination of the boundaries of the BOLD fMRI maps. The averaged BOLD fMRI signal was distributed unevenly over the extent of the map and the data at the boundaries could be modeled with major and minor spatial components. Comparison of three-dimensional echo-planar imaging (EPI) fMRI at isotropic 300 µm resolution demonstrated that the border locations of the major spatial component of BOLD signal did not overlap between the forepaw and hindpaw maps. Interestingly, the border positions of the minor BOLD fMRI spatial components extended significantly into neighboring representations. Similar results were found for cerebral blood volume (CBV) weighted fMRI obtained using iron oxide particles, suggesting that the minor spatial components may not be due to vascular mislocalization typically associated with BOLD fMRI. Comparison of the BOLD fMRI maps of the forepaw and hindpaw to histological determination of these representations using cytochrome oxidase (CO) staining demonstrated that the major spatial component of the BOLD fMRI activation maps accurately localizes the borders. Finally, 2-3 weeks following peripheral nerve denervation, cortical reorganization/plasticity at the boundaries of somatosensory limb representations in adult rat brain was studied. Denervation of the hindpaw caused a growth in the major component of forepaw representation into the adjacent border of hindpaw representation, such that fitting to two components no longer led to a better fit as compared to using one major component. The border of the representation after plasticity was the same as the border of its minor component in the absence of any plasticity. It is possible that the minor components represent either vascular effects that extend from the real neuronal representations or the neuronal communication between neighboring regions. Either way the results will be useful for studying mechanisms of plasticity that cause alterations in the boundaries of neuronal representations.

3D mapping of somatotopic reorganization with small animal functional MRI.

There are few in vivo noninvasive methods to study neuroplasticity in animal brains. Functional MRI (fMRI) has been developed for animal brain mapping, but few fMRI studies have analyzed functional alteration due to plasticity in animal models. One major limitation is that fMRI maps are characterized by statistical parametric mapping making the apparent boundary dependent on the statistical threshold used. Here, we developed a method to characterize the location of center-of-mass in fMRI maps that is shown not to be sensitive to statistical threshold. Utilizing centers-of-mass as anchor points to fit the spatial distribution of the BOLD response enabled quantitative group analysis of altered boundaries of functional somatosensory maps. This approach was used to study cortical reorganization in the rat primary somatosensory cortex (S1) after sensory deprivation to the barrel cortex by follicle ablation (F.A.). FMRI demonstrated an enlarged nose S1 representation in the 3D somatotopic functional maps. This result clearly demonstrates that fMRI enables the spatial mapping of functional changes that can characterize multiple regions of S1 cortex and still be sensitive to changes due to plasticity.

Brain redox imaging using blood-brain barrier-permeable nitroxide MRI contrast agent.

Reactive oxygen species (ROS) and compromised antioxidant defense may contribute to brain disorders such as stroke, amyotrophic lateral sclerosis, etc. Nitroxides are redox-sensitive paramagnetic contrast agents and antioxidants. The ability of a blood-brain barrier (BBB)-permeable nitroxide, methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl (MC-P), as a magnetic resonance-imaging (MRI) contrast agent for brain tissue redox imaging was tested. MC-P relaxation in rodent brain was quantified by MRI using a fast Look-Locker T(1)-mapping sequence. In the cerebral cortex and thalamus, the MRI signal intensity increased up to 50% after MC-P injection, but increased only by 2.7% when a BBB-impermeable nitroxide, 3CxP (3-carboxy-2,2,5,5,5-tetramethylpyrrolidine-1-oxyl) was used. The maximum concentrations in the thalamus and cerebral cortex after MC-P injection were calculated to be 1.9+/-0.35 and 3.0+/-0.50 mmol/L, respectively. These values were consistent with the ex vivo data of brain tissue and blood concentration obtained by electron paramagnetic resonance (EPR) spectroscopy. Also, reduction rates of MC-P were significantly decreased after reperfusion following transient MCAO (middle cerebral artery occlusion), a condition associated with changes in redox status resulting from oxidative damage. These results show the use of BBB-permeable nitroxides as MRI contrast agents and antioxidants to evaluate the role of ROS in neurologic diseases.

BOLD fMRI and somatosensory evoked potentials are well correlated over a broad range of frequency content of somatosensory stimulation of the rat forepaw.

Electrical stimulation of the rat paw is commonly used to study the hemodynamic, metabolic and neuronal mechanisms of functional MRI (fMRI) responses in somatosensory cortex. Several groups have reported good correlation between the blood oxygenation level-dependent (BOLD) fMRI signal and somatosensory evoked potentials (SEPs) using short, typically 300 micros, square stimulation pulses. The spectral power of these short pulses is evenly distributed over a wide range of frequencies and thus the effects of the frequency content of the stimulation pulse on fMRI responses have not been previously described. Here, the effects that different stimulation pulse waveforms with a range of frequency content have on neuronal activity, as measured by SEPs, and on the amplitude of the BOLD fMRI signal in rat somatosensory cortex are investigated. The peak-to-peak SEP amplitudes increased as the power in the high frequency harmonics of the different pulse waveforms increased, using either triangular or sinusoidal stimuli waveforms from 9 Hz to 180 Hz. Similarly, BOLD fMRI response increased with increased high frequency content of the stimulation pulse. There was a linear correlation between SEPs and BOLD fMRI over the full range of frequency content in the stimulations.

Transport and imaging of brute-force (13)C hyperpolarization.

We demonstrate transport of hyperpolarized frozen 1-(13)C pyruvic acid from its site of production to a nearby facility, where a time series of (13)C images was acquired from the aqueous dissolution product. Transportability is tied to the hyperpolarization (HP) method we employ, which omits radical electron species used in other approaches that would otherwise relax away the HP before reaching the imaging center. In particular, we attained (13)C HP by 'brute-force', i.e., using only low temperature and high-field (e.g., T<∼2K and B∼14T) to pre-polarize protons to a large Boltzmann value (∼0.4% (1)H polarization). After polarizing the neat, frozen sample, ejection quickly (<1s) passed it through a low field (B<100G) to establish the (1)H pre-polarization spin temperature on (13)C via the process known as low-field thermal mixing (yielding ∼0.1% (13)C polarization). By avoiding polarization agents (a.k.a. relaxation agents) that are needed to hyperpolarize by the competing method of dissolution dynamic nuclear polarization (d-DNP), the (13)C relaxation time was sufficient to transport the sample for ∼10min before finally dissolving in warm water and obtaining a (13)C image of the hyperpolarized, dilute, aqueous product (∼0.01% (13)C polarization, a >100-fold gain over thermal signals in the 1T scanner). An annealing step, prior to polarizing the sample, was also key for increasing T1∼30-fold during transport. In that time, HP was maintained using only modest cryogenics and field (T∼60K and B=1.3T), for T1((13)C) near 5min. Much greater time and distance (with much smaller losses) may be covered using more-complete annealing and only slight improvements on transport conditions (e.g., yielding T1∼5h at 30K, 2T), whereas even intercity transfer is possible (T1>20h) at reasonable conditions of 6K and 2T. Finally, it is possible to increase the overall enhancement near d-DNP levels (i.e., 10(2)-fold more) by polarizing below 100mK, where nanoparticle agents are known to hasten T1 buildup by 100-fold, and to yield very little impact on T1 losses at temperatures relevant to transport.