Bock, Christian

Brinker, G.

Hoehn-Berlage, M.

Krep, H.

English

Electronic Publication Information Center

Brainmapping of a-chloralose anesthetized rats withT2*-weighted imaging: distinction between therepresentation of the forepaw and hindpaw in thesomatosensory cortex†C. Bock, H. Krep, G. Brinker, M. Hoehn-Berlage*Max-Plank-Institute for Neurological Research, Cologne, GermanyReceived 26 September 1997; revised 7 January 1998; accepted 7 January 1998ABSTRACT: T2*-weighted imaging at 4.7 T was used to identify the cortical areas activated by electrical stimulationof the forepaw and hindpaw of a-chloralose anesthetized rats. Variation of the coronal slice position relative to thebregma, showed that the forepaw representation in the somatosensory cortex is more frontal and lateral than that ofthe hindpaw. Overlap between both activation areas was observed only in a small region in the slice at the level of thebregma. Documented localizations of both representations are in good agreement with earlier observations usinginvasive techniques. The determination of the separate areas of both paws indicates the feasibility of more complexactivation studies in anesthetized animals, such as combined stimulations for the investigation of potentiation ordepression effects on individual stimuli. Ó 1998 John Wiley & Sons, Ltd.KEYWORDS: fMRI; forepaw; hindpaw; rat; somatosensory cortexINTRODUCTIONThe application of functional magnetic resonanceimaging (fMRI) to animal models has recently beendemonstrated in various publications.1–4 In most of thesestudies somatosensory activation was performed withelectrical forepaw stimulation of a-chloralose anesthe-tized rats. Recently, fMRI studies with this animal modelwere extended to investigations under pathophysiologicalconditions.5–7 There, variations of the fMRI signalintensity were observed depending on the differentphysiological or pathophysiological conditions5–7 orstimulation paradigm.2,8 All these studies were limitedto somatosensory activation of the forepaw with varia-tions from left to right forepaw, or right and leftsomatosensory cortex, respectively.2,3 The first reportdescribing the activation of neighboring brain areas in thesomatosensory cortex of the rat with fMRI was presentedby Yang et al.9 These authors were able to detectdifferent activated regions in the somatosensory cortexduring single barrel stimulation at 7 T.In the present study, we demonstrate the feasibility todistinguish between different activated regions in thesomatosensory cortex using fore- and hindpaw stimula-tions at 4.7 T. The cortical representations of both pawshave been described to be direct neighbors to eachother.10 Together, they make up a large portion of thesomatosensory cortex, S1. Separation of the activatedregions in the somatosensory cortex will provide the basisfor more complex stimulation paradigms in animalsunder different physiological conditions, such as inter-actions of stimuli or enlargement of representation areaafter conditioning.11MATERIALS AND METHODSAnimal modelFive Sprague-Dawley rats were initially anesthetizedwith 1.5% halothane in a gas mixture of 70/30 v/v N2O/O2 for surgical preparation. The femoral artery and veinwere catheterized with PE50 tubing for blood pressuremonitoring, fluid and drug administration, and bloodsampling. After a tracheotomy, the animals wereparalyzed with pancuronium bromide (0.1 mg/kg/h) andNMR IN BIOMEDICINENMR Biomed 11, 115–119 (1998)*Correspondence to: Dr Mathias Hoehn-Barlage, In-vivo-NMR, Max-Planck-Institut fu¨r neurologische Forschung, Gleuelerstr. 50, D-50931Ko¨ln, Germany. Tel.: (49) 221-4726-315 Fax.: (49) 221-4726-298.Email: mathias@mpin-koeln.mpg.de†Abbreviations used: fMRI, functional magnetic resonance imaging;T2*-WI, T2*-weighted imaging.Contract/grant sponsor: Deutshe Forschungsgemeinschaft; contractgrant number: SFB194/B1.Ó 1998 John Wiley & Sons, Ltd. CCC 0952–3480/98/030115–05 $17.50artificially ventilated with a small animal ventilator(RVS/1301, FMI-Medical Instruments, Seeheim, Ger-many). Then, anesthesia was switched to a-chloralose(80 mg/kg i.v.) for functional activation studies asalready described in detail by Gyngell et al.2 and Uekiet al.12 Body temperature was measured via a rectalthermocouple and kept at 37 ° C with a feedback-controlled warm water blanket. Arterial blood pressurewas monitored continuously, and arterial blood samplesfor blood gas analysis were taken in 30 min intervals.For somatosensory stimulation, pairs of small needleelectrodes were inserted into the skin of the left or rightfore- and hindpaw. Electrical stimulation of the paws wascarried out by applying square-wave pulses of 0.3 msduration with a frequency of 3 Hz.2 The current was 0.5mA, unless stated otherwise.NMR methodsThe NMR measurements were performed at 4.7 T using a47/30 Biospec MSL-X11 system (Bruker Medizintech-nik, Ettlingen, Germany) with actively shielded gradientcoils (100 mT/m, 250 ms rise-time). Radio frequencypulses were transmitted using a 12 cm diameter Helm-holtz coil, with the NMR signal reception via a 16 mmdiameter inductively coupled surface coil placed over theskull and centered over the midline of the animal. Multi-slice FLASH pilot scans of brain anatomy were obtainedin the sagittal planes, in order to place the functionalimaging slice coronally through the fore- or hindpaw-area of the somatosensory cortex choosing the positionrelative to the rhinal fissure. For functional studies T2*-weighted images (T2*-WI) were acquired using a FLASHsequence (TR = 70 ms, TE = 60 ms, matrix size =64 64, FOV = 2.4 cm, 2 mm slice thickness) with aflip angle a of 22.5° , resulting in a scan time of 4.5 s.2Anatomical coronal FLASH images of every fMRI slice(TR = 400 ms, TE = 8.4 ms, matrix size = 128 128,FOV = 2.4 cm, 2 mm slice thickness) were recorded forcomparison with anatomical data from the literature byPaxinos10 and Zilles,13 and for data analysis.Experimental protocolThe animals were fixed in a non-magnetic stereotacticheadholder and positioned in the magnet. A set of eightT2* -WIs were collected prior to stimulation (baseline)and were repeated during stimulation. All experimentsstarted with the electrical stimulation of the forepaw inorder to test the functional response of the somatosensorycortex in the T2*-weighted images, and for comparisonwith established protocols of previous investigations.2,3,7After a resting period of 5 min electrical stimulation ofthe hindpaw was carried out. Then, the position of theimaging slice was shifted 1 mm posterior or anteriorrelative to the bregma and the set was repeated. In somecases, stimulation of the hindpaw was repeated as afunction of the electrical current (I = 0.5, 0.75, 1.0, 1.25and 2.0 mA).T2*-WI activation maps were obtained by subtractingaveraged baseline images from averaged activationimages, taking only those pixels into account with signalintensity 1.5 standard deviations (SD) above backgroundnoise.2 Pixels were included only inside the brain andwhen forming spatial clusters of at least three pixels.Resulting activation maps were overlaid on anatomicalFLASH images of the corresponding slice.RESULTSThe physiological parameters remained within thenormal range during the whole length of the NMRexperiments. Blood pressure was kept between 90 and100 mmHg, and arterial pCO2 between 35 and 40 mmHg.Figure 1 presents a schematic drawing of the localiza-tion of different somatosensory regions in the rat brain,based on the regional assignment by Paxinos.10 The areaof the forepaw is located more anterior and lateral in thesomatosensory cortex than that of the hindpaw. Fororientation purposes, the coronal image positions chosenfor the activation experiments of fore- and hindpaw arealso included in the drawing.Stimulation of the right forepaw with a current of 0.5mA resulted in a clear increase in T2*-WI signal intensityFigure 1. Schematic drawing of an axial orientation of therat brain from the dorsal view. The olfactory bulb (OB) andthe rhinal ®ssure (RF) are marked. The representations of theright forepaw (FP) and hindpaw (HP) in the somatosensorycortex of the left hemisphere are circumscribed. Fororientation of the image positions of the fMRI experiments,the four chosen coronal slices are indicated. For localizationof the image sections, their center positions are given relativeto the bregma. The bregma (BG) is marked on the midline.Ó 1998 John Wiley & Sons, Ltd. NMR IN BIOMEDICINE, VOL. 11, 115–119 (1998)116 C. BOCK ET AL.in the somatosensory cortex of the left hemisphere (Fig.1). In contrast, the same stimulation of the hindpaw didnot lead to any signal intensity changes of the T2*-WI.Only when raising the current to 1.25 mA during hindpawstimulation was an activated area detected. No significantdifference in T2*-WI signal intensity increase wasobserved between the activation of fore- and hindpaw.An increase in blood pressure was not observed at thiscurrent during hindpaw stimulation, in contrast to theforepaw where the blood pressure increased when thestimulation current was raised above 1 mA.In Fig. 2 the functional activation maps of differentbrain slices during the stimulation of the right fore- andhindpaw, respectively, are presented. On the left columnfMRIs during forepaw stimulation are shown (I = 0.5mA); activation maps during hindpaw stimulation(I = 1.25 mA) are given in the center column of Fig. 2.The corresponding positions of the image planes relativeto the bregma are indicated on the right.While a highlighted area in the left somatosensorycortex during forepaw stimulation occured in the mostanterior slice (2 mm relative to the bregma) andincreased in size in the slice 1 mm anterior to the bregma,no activation was observed in these slices during hindpawstimulation. In the following slice (at the bregma),posterior to the former two slices, the size of the activatedarea of the forepaw stimulation decreased again in the leftsomatosensory cortex. Stimulation of the hindpawresulted in a small activated area in the somatosensorycortex of the same slice. This activated region wasFigure 2. Coronal images of the rat brain at various positions anterior to posterior of the bregma. The activated areas, analyzedfrom the T2*-weighted image experiments, are overlaid on anatomical FLASH images. Activation of the forepaw is seen on thetwo frontal sections with the activation center being in the slice 1 mm anterior of the bregma. The activated area duringhindpaw stimulation appears only when the imaged sections are shifted further posterior. Also, the hindpaw representation ismore medial than that of the forepaw.Ó 1998 John Wiley & Sons, Ltd. NMR IN BIOMEDICINE, VOL. 11, 115–119 (1998)fMRI OF FOREPAIN AND HINDPAN ACTIVATION IN RAT 117located slightly more medial than for the forepaw. Whenthe image plane was moved 1 mm posterior to thebregma, the activation during forepaw stimulation was nolonger observed in the left somatosensory cortex,whereas the activated area during hindpaw stimulationincreased in size.Shifting the image plane even further posterior to thebregma, the T2*-weighted images showed strong arti-facts, because of susceptibility effects of the air-filledpetrous bones of the animals. Comparison of the positionof the activated regions as found from our fMRI dataagreed very well with the assigment as described byPaxinos10 (Fig. 1).DISCUSSIONThe aim of this paper was to demonstrate the capability offMRI to achieve successful brainmapping in anesthetizedanimals at 4.7 T. The schematic functional architecture ofthe rat brain shows two separate, but closely neighboring,areas for the representation of the forepaw and hindpawin the somatosensory cortex.10 The locations of theseareas differ in their position anterior and posteriorrelative to the bregma, but there is also a coronal sectionwhere both regions can be detected simultaneously.Furthermore, the shift of the activated areas from lateralto medial when going from fore- to hindpaw stimulation,was presented in the T2*-WI activation maps of Fig. 2.Also, a small overlap of both regions could be observedin the T2*-WI activation maps in the image plane at thebregma position (0 mm), indicating the good spatialresolution of fMRI at 4.7 T.It should be pointed out that the stimulation currents ofthe activation paradigms of fore- and hindpaw weredifferent. This is explained with the differing sensitivityof fore- and hindpaw to somatosensory stimulation. Inman, the hands (and fingers, in particular) are known tohave the highest somatosensory receptor density. Similarto that situation, the forepaw of the rat may be consideredhighly sensitive to somatosensory stimulations becausethese animals use their forepaws for all somatosensoryactivities, whereas the hindpaws are used predominantlyfor standing and running. Therefore, it is conceivable thatthe hindpaws require a higher electrical current foractivation. Owing to the high sensitivity of the forepaw, astimulation current above 0.5 mA induced a rise in bloodpressure. This was in contrast to hindpaw stimulation,where no increase of blood pressure was observed. Onlywhen the stimulation current was raised further to 2 mA,was an increase of blood pressure induced duringhindpaw stimulation (data not shown), confirming thelesser sensitivity of the hindpaw to electrical stimulation.These differences indicate that the control of stimulationparadigms for functional activation studies in animals areimportant in order to avoid a misinterpretation of fMRIdata, particularly for more complex stimulation proto-cols.CONCLUSIONSStimulation of the fore- and hindpaw resulted in differentactivated areas in the somatosensory cortex of the ratbrain observed with fMRI, demonstrating the capabilityof T2*-weighted imaging and its sufficient spatialresolution for brainmapping studies in the rat brain at4.7 T. Furthermore, the different stimulation currents foractivation of fore- and hindpaw, indicated the necessity tocontrol and/or adapt the stimulation paradigms forparticular applications. The possibility for fMRI todistinguish different activated regions in the rat brain at4.7 T prepares the ground for investigations of combinedand more complex stimulation paradigms, including theresult of somatosensory interactions between individualstimuli. Such experiments are at present under way in ourlaboratory.AcknowledgementThe study was supported in part by a grant from theDeutsche Forschungsgemeinschaft (SFB194/B1).REFERENCES1. Hyder F., Behar K. L., Martin M. A., Blamire A. M. and ShulmanR. G. Dynamic magnetic resonance imaging of the rat brain duringforepaw stimulation. J. Cereb. Blood Flow Metab. 14, 649–655(1994).2. Gyngell M. L., Bock C., Schmitz B., Hoehn-Berlage M. andHossmann K.-A. Variation of functional MRI signal in response tofrequency of somatosensory stimulation in alpha-chloraloseanesthetized rats. Magn. Reson. Med. 36, 13–15 (1996).3. Kerskens C. M., Hoehn-Berlage M., Schmitz B., Busch E., BockC., Gyngell M. L. and Hossmann K.-A. Ultrafast perfusion-weighted MRI of functional brain activation in rats during forepawstimulation. Comparison withT2*-weighted MRI. NMR Biomed. 8,20–23 (1996).4. Huang W., Palyka I., Li H.-F., Eisenstein E. M., Volkow N. D. andSpringer Jr C. S. Magnetic resonance imaging (MRI) detection ofthe murine brain response to light: temporal differentiation andnegative functional MRI changes. Proc. Natl Acad. Sci. USA 93,6037–6042 (1996).5. Hossmann K.-A., Schmitz B., Bock C., Kerskens C. and Hoehn-Berlage M. Functional magnetic resonance imaging after cardiacarrest in rat. In Pharmacology of Cerebral Ischemia, pp. 357–365.Medpharm Scientific Publishers, Stuttgart (1996).6. Schmitz B., Bock C., Kerskens C., Hoehn-Berlage M., Bo¨ttiger B.W. and Hossmann K.-A. Recovery of the coupling of cerebralblood flow to neuronal activation following cardiac arrest in rats: afunctional fMRI study. Int. Soc. Magn. Reson. 440 (1996) Book ofabstracts.7. Bock C., Schmitz B., Kerskens C. M., Gyngell M. L., HossmannK.-A. and Hoehn-Berlage M. Functional MRI of somatosensoryactivation in rat: effect of hypercapnic upregulation on perfusion-and BOLD-imaging. Magn. Reson. Med. 39, 457–461 (1998).8. Scanley B. E., Kennan R. P., Cannan S., Skudarski P., Innis R. B.and Gore J. C. Functional magnetic resonance imaging of medianÓ 1998 John Wiley & Sons, Ltd. NMR IN BIOMEDICINE, VOL. 11, 115–119 (1998)118 C. BOCK ET AL.nerve stimulation in rats at 2.0 T. Magn. Reson. Med. 37, 969–972(1997).9. Yang X., Hyder F. and Shulman R. G. Activation of single whiskerbarrel in rat brain localized by functional magnetic resonanceimaging. Proc. Natl Acad. Sci. USA 93, 475–478 (1996).10. Paxinos G. The Rat Nervous System. Academic Press, San Diego(1995).11. Godde B., Spengler F. and Dinse H. R. Associative pairing oftactile stimulation induces somatosensory cortical reorganizationin rats and humans. NeuroRep. 8, 281–285 (1996).12. Ueki M., Linn F. and Hossmann K.-A. Functional activation ofcerebral blood flow and metabolism before and after globalischaemia of rat brain. J. Cereb. Blood Flow Metab. 8, 486–494(1988).13. Zilles K. The Cortex of the Rat: A Stereotactic Atlas. Springer,Berlin, (1985).Ó 1998 John Wiley & Sons, Ltd. NMR IN BIOMEDICINE, VOL. 11, 115–119 (1998)fMRI OF FOREPAIN AND HINDPAN ACTIVATION IN RAT 119

Brainmapping of a-chloralose anesthetized rats with T2*-weighted imaging-distinction between the representation of the forepaw and hindpaw in the somatosensory cortex

Brainmapping of a-chloralose anesthetized rats with T2*-weighted imaging-distinction between the representation of the forepaw and hindpaw in the somatosensory cortex

Abstract

Similar works

Full text

Available Versions

Electronic Publication Information Center