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Prolonged anoxic depolarization exacerbates NADH hyperoxidation and promotes poor electrical recovery after anoxia in hippocampal slices.

Mitochondrial dysfunction appears to occur during brain ischemia and following reperfusion. A characteristic event during reoxygenation after anoxia in hippocampal slices is hyperoxidation of the electron carriers of the mitochondrial respiratory chain. Earlier studies suggested that calcium influx due to loss of ion homeostasis during anoxia was linked to neuronal damage. Since a link between cytosolic calcium overload and post-anoxic hyperoxidation (PAMHo) has been suggested in previous studies, present studies sought to test the hypothesis that the length of anoxic depolarization can influence hyperoxidation and electrical activity recovery following anoxia in hippocampal slices. Rat hippocampal slices were made anoxic and then allowed to recover for 60 min. The time of anoxia was defined by the time of anoxic depolarization (AD), and slices were divided in five groups: 0.5, 1, 2, 5 and 10 min of AD. Reduction/oxidation shifts of NADH were measured by rapid scanning spectrofluorometry. Synaptic activity was indicated by population spike amplitudes in the CA1 pyramidal cell subfield of the hippocampus in response to stimulation of the Schaffer collaterals. We report here that mitochondrial hyperoxidation and synaptic activity in hippocampal slices are highly sensitive to the time in which slices remain depolarized (AD).

Animals↗

Imaging of gene expression during long-term potentiation.

Long term potentiation (LTP) at hippocampal Schaffer collateral-CA1 synapses involves an early and a late phase, where only the latter is sensitive to protein synthesis inhibitors. Here we characterized the dynamics of protein synthesis associated with the induction of L-LTP using a transgenic mouse model in which a cAMP responsive element (CRE)-regulated promoter drives production of an enhanced yellow fluorescent protein (eYFP). We found that eYFP fluorescence increased after less than 30 min following L-LTP induction. Application of transcription and translation suppressors and the NMDA receptor antagonist D-AP5 inhibited the L-LTP and prevented the rise in eYFP levels. The early-phase of LTP was not affected by inhibiting protein synthesis.

2-Amino-5-phosphonovalerate↗

A clinically relevant small-animal model of skeletal muscle ischemia-reperfusion injury.

Animal models of skeletal muscle ischemia-reperfusion injury have led to a better understanding of the pathophysiology of this condition and are necessary for the evaluation of potential therapeutics. This study presents a new, well-controlled model of ischemia-reperfusion that more accurately simulates acute arterial occlusions in humans. In rabbits, a whole hindlimb is rendered reversibly ischemic by occlusion of primary and collateral arterial inflow and then reperfused for an extended period of 48 h. Ischemic injuries are standardized by defining and controlling ischemic time, limb temperature, and the extent of collateral circulation. We have characterized this model by measuring anterior tibial and soleus muscle necrosis and edema formation in groups of animals subjected to 4 h of ischemia at either 32 or 36 degrees C, with one of two extents of collateralization and with or without muscle compartment release (fasciotomy). Our results indicate the following: (1) muscle necrosis is significantly worsened by restricting the extent of collateral blood supply or by elevating ischemic temperature; (2) anterior tibial muscle is inherently more sensitive than soleus muscle to ischemic injury; (3) fasciotomy may reduce muscle necrosis by more than 50%; and (4) the amount of edema present in muscles is an unreliable indicator of actual muscle necrosis. We conclude that this new model is a practical, well-controlled, and clinically relevant preparation useful for the investigation of ischemic muscle injury.

Animals↗

Different modes of action of substance P in the motor control of the feline stomach and pylorus.

In an experimental in vivo model to study gastropyloric motility in the cat a contraction of the stomach and the pyloric sphincter was regularly obtained in animals subjected to electrical vagal nerve stimulation or local intraarterial (i.a.) injection of substance P (SP). Much more infrequently contractile motor responses were recorded at splanchnic nerve stimulation. The contractile effects of SP were sensitive to atropine or local infusion of a SP analogue, (D-Pro2,D-Trp7,9)-SP, indicating that SP activated a final common cholinergic neuron in both stomach and pylorus. However, there seemed to be separate transmission mechanisms in these two regions based on the results of the physiological studies. The vagally induced pyloric contraction was noncholinergic, nonadrenergic, but sensitive to ganglionic blockade (hexamethonium) or the SP analogue, indicating involvement of SP in a peptidergic pathway to the sphincter. The infrequent splanchnically induced pyloric contraction was sensitive to atropine, the SP analogue or ganglionic blockade (hexamethonium) in favour of SP acting on a final cholinergic neuron in this system. On the other hand the gastric contraction, obtained at either extrinsic nerve stimulations or local i.a. injection of SP, was sensitive to atropine or the SP analogue but hexamethonium resistant. These findings suggest antidromic activation of SP-containing axon collaterals of the extrinsic nerves terminating on cholinergic neurons of the gastric wall. When afferent C-fibres of the vagal nerve were selectively activated by local heating, pyloric contraction and gastric relaxation were obtained via vago-vagal reflexes. After cervical vagotomy heating of the distal end of the vagal nerve elicited a gastric contraction, previously demonstrated to be atropine sensitive and hexamethonium resistant, but no pyloric motor response. This suggests that the antidromic activation mechanism was present only in the stomach, not in the pylorus.

Animals↗

Transcutaneous PO2: a potential monitor the status of replanted limb parts.

In animal and human investigations, transcutaneous PO2 proved to be a sensitive and rapid indicator of the patency of the artery and veins supplying a limb part. Furthermore, transcutaneous PO2 indicated the presence or absence of collateral circulation. Commercially available transcutaneous PO2 probes were easily applied to human fingers and dramatically reflected the compromise of finger circulation by the inflation of a blood-pressure cuff around the arm. Probe temperatures from 42 to 44 degrees C appeared to provide satisfactory sensitivity and safety. On the basis of these studies, we conclude that transcutaneous PO2 monitoring holds sufficient potential for the evaluation of replanted limb parts to deserve a clinical trial.

Animals↗

Characterization of the mechanical behavior of human knee ligaments: a numerical-experimental approach.

During knee-joint motions, the fiber bundles of the knee ligaments are nonuniformly loaded in a recruitment pattern, which depends on successive relative orientations of the insertion sites. These fiber bundles vary with respect to length, orientation and mechanical properties. As a result, the stiffness characteristics of the ligaments as a whole are variable during knee-joint motion. The purpose of the present study is to characterize this variable mechanical behavior. It is hypothesized that for this purpose it is essential to consider the ligaments mechanically as multi-bundle structures in which the variability in fiber bundle characteristics is accounted for, rather than as one-dimensional structures. To verify this hypothesis, bone-ligament-bone preparations of the ligaments were subjected to series of unidirectional subfailure tensile tests in which the relative insertion orientations were varied. For each individual test specimen, this series of tensile tests was simulated with a mathematical ligament model. Geometrically, this model consists of multiple line elements, of which the insertions and orientations are anatomically based. In a mathematical optimization process, the unknown stiffness and recruitment parameters of the line elements are identified by fitting the variable stiffness characteristics of the model to those of the test series. Thus, lumped parameters are obtained which describe the mechanical behavior of the ligament as a function of the relative insertion orientation. This method of identification was applied to all four knee ligaments. In all cases, a satisfactory fit between experimental results and computer simulation was obtained, although the residual errors were lower for the cruciate ligaments (1.0-2.4%) than for the collateral ligaments (3.7-8.1%). It was found that models with three or less line elements were very sensitive to geometrical parameters, whereas models with more than 7 line elements suffered from mathematical redundancy. Between 4 and 7 line elements little difference was found. It is concluded that the present ligament models can realistically simulate the variable tensile behavior of human knee ligaments. Hereby the hypothesis is verified that it is essential to consider the ligaments of the knee as multi-bundle structures in order to characterize fully their mechanical behavior.

Aged↗

Continuous ST-segment monitoring during coronary angioplasty using orthogonal ECG leads.

In order to characterize ST-segment shifts during transient coronary artery occlusion, 24 patients with single-vessel disease were continuously monitored during percutaneous transluminal coronary angioplasty by use of a computerized orthogonal lead system. Changes of ST-segment (J + 60 ms) in leads X, Y, and Z and of the ST vector magnitude were analyzed by using 20 microV as a threshold for significant ST-segment shift. The sensitivity and magnitude of this shift were compared among the left anterior descending, right coronary, and circumflex artery groups (11, 8, and 5 patients, respectively) during balloon inflation. Significant ST-segment shifts were seen in 22 patients (92%) in ST-VM, Y, and Z leads and all patients in lead X (100%). There was no significant difference in sensitivity of either the ST vector magnitude or the most sensitive lead for occlusion detection among the three groups. There was a significantly greater magnitude of ST shift during left anterior descending artery occlusion than during right coronary artery and circumflex artery occlusions in ST-VM. Analysis of the direction of ST shifts in the X, Y, and Z leads showed a characteristic pattern, which could distinguish among the three coronary groups in 21 patients (88%). The presence of collaterals was significantly associated with ST-segment depression in leads oriented toward ischemia (3 of 6 patients) as compared with ST-segment elevation in the absence of collaterals (all of 15 patients), P > .01. It is concluded that ST-segment shift in the orthogonal leads is a reliable marker for myocardial ischemia. It is equally sensitive to occlusion of each of the three major coronary arteries and can thus identify the occluded coronary. An ST-segment depression instead of an elevation was related to the presence of collaterals, which may reflect a lesser degree of ischemia.

Adult↗

Hepatic hemangioma in cirrhotics with portal hypertension: evaluation with Tc-99m red blood cell SPECT.

PURPOSE: To determine whether extensive shunting associated with portal hypertension (PH) adversely affects the diagnostic accuracy of technetium-99m red blood cell (RBC) single photon emission computed tomography (SPECT) for hepatic hemangioma (HH). MATERIALS AND METHODS: Nineteen possible HHs (1.0-11.0 cm in diameter) in 15 cirrhotics with PH were studied with RBC SPECT. RESULTS: RBC SPECT allowed correct categorization of four HHs (1.5, 2.8, 4.0, and 4.0 cm in diameter) and 13 non-HHs (1.5-11.0 cm in diameter). Two small HHs (1.0 and 1.4 cm in diameter) were "missed." Sensitivity, specificity, and accuracy were 67%, 100%, and 89%, respectively. CONCLUSION: Despite abnormal liver blood flow dynamics due to portosystemic collateralization, RBC SPECT allows reliable identification of HH in cirrhotics with PH. As in noncirrhotics, its major limitation is small lesion size.

Adult↗

Peripheral and central predictors of whisker afferent morphology in the rat brainstem.

Prior studies suggest that whisker afferents have but one central projection pattern, despite their association with differing peripheral receptors that predict central morphology in other systems. Target factors in barrelettes are thought to dictate afferent projection patterns; yet, barrelettes differ in their size, shape and development. We tested the hypothesis that whisker afferents have differing morphologies that are predicted by peripheral and central factors. Branching patterns and collaterals of 78 Neurobiotin-stained afferents were compared in rats. Fibers from one whisker had precisely somatotopic projections but highly varied morphologies. For the entire sample, analysis of variance revealed significant intrafiber variance in collateral number and arbor shape that was attributed to the target subnucleus. Significant interfiber variance did not reflect response adaptation rate, direction sensitivity, whisker row origin or parent fiber bifurcation in the trigeminal root. Instead, we found the following. 1) Mandibular fibers had more elongated arbors than maxillary axons. In subnuclei interpolaris and principalis, mandibular fibers had larger arbors with more boutons/collateral than maxillary axons; in oralis and interpolaris, mandibular fibers had fewer collaterals than those of the maxillary division. 2) Upper lip whisker axons had more boutons than those from the B-D row in all subnuclei. 3) Rostral whisker are afferents had larger arbors and more boutons than those from middle or caudal arcs due to significant arc effects in interpolaris and oralis. Thus, whisker afferents are not structurally uniform, and some morphological features are predictable. Intrafiber variance is attributed to the central target; interfiber variance reflects maxillary versus mandibular origin, upper lip origin and whisker rostrocaudal arc.

Adaptation, Physiological↗

Relationship between contractile reserve, Tl-201 uptake, and collateral angiographic circulation in collateral-dependent myocardium: implications regarding the evaluation of myocardial viability.

BACKGROUND: The aim of this study was to compare collateral angiographic circulation (CC), thallium 201 single photon emission computed tomography (SPECT), low-dose dobutamine echocardiography (LDDE), and their combination in identifying viable myocardium beyond a chronic occlusion. METHODS AND RESULTS: We studied 33 consecutive patients with a chronic occluded coronary artery and regional ventricular dysfunction who underwent dobutamine infusion (5-10 microg x kg(-1) x min(-1)), Tl-201 SPECT, and coronary revascularization (15 patients underwent coronary artery bypass grafting and 18 patients percutaneous transluminal coronary angioplasty). Echocardiography and Tl-201 SPECT at rest were repeated 90 +/- 48 days after revascularization. For viability assessment, Tl-201 SPECT showed the best sensitivity and accuracy (77% and 72%, respectively) compared with LDDE (64% and 63%, respectively) and CC (74% and 55%, P < .05 vs Tl-201 SPECT). Specificity was significantly better for both Tl-201 SPECT (65%) and LDDE (62%) compared with collateral circulation (27%, P < .001 vs Tl-201 SPECT and LDDE). Furthermore, combined Tl-201 SPECT and LDDE data improved specificity (from 88% to 92%) and positive predictive power (from 75% to 78%) but not global accuracy. A direct relationship between the number of viable segments and global functional recovery after revascularization was found only in the case of Tl-201 SPECT (r = 0.48, P = .04). CONCLUSIONS: This study demonstrates that CC has a very low specificity in the evaluation of viability in collateral-dependent myocardium and that LDDE has a very low sensitivity, whereas Tl-201 SPECT is the most reliable method, particularly in akinetic segments, and optimally identifies patients who will mostly benefit from revascularization in terms of ventricular functional improvement. No additional advantages in terms of accuracy were found when Tl-201 SPECT and LDDE data were combined.

Aged↗

Exercise training restores adenosine-induced relaxation in coronary arteries distal to chronic occlusion.

We previously reported that canine collateral-dependent coronary arteries exhibit impaired relaxation to adenosine but not sodium nitroprusside. In contrast, exercise training enhances adenosine sensitivity of normal porcine coronary arteries. These results stimulated the hypothesis that chronic coronary occlusion and exercise training produce differential effects on cAMP- versus cGMP-mediated relaxation. To test this hypothesis, Ameroid occluders were surgically placed around the proximal left circumflex coronary artery (LCx) of female Yucatan miniature swine 8 wk before initiating sedentary or exercise training (treadmill run, 16 wk) protocols. Relaxation to the cAMP-dependent vasodilators adenosine (10(-7) to 10(-3) M) and isoproterenol (3 x 10(-8) to 3 x 10(-5) M) were impaired in collateral-dependent LCx versus nonoccluded left anterior descending (LAD) arterial rings isolated from sedentary but not exercise-trained pigs. Furthermore, adenosine-mediated reductions in simultaneous tension and myoplasmic free Ca(2+) were impaired in LCx versus LAD arteries isolated from sedentary but not exercise-trained pigs. In contrast, relaxation in response to the cAMP-dependent vasodilator forskolin (10(-9) to 10(-5) M) and the cGMP-dependent vasodilator sodium nitroprusside (10(-9) to 10(-4) M) was not different in LCx versus LAD arteries of sedentary or exercise-trained animals. These data suggest that chronic occlusion impairs receptor-dependent, cAMP-mediated relaxation; receptor-independent cAMP- and cGMP-mediated relaxation were unimpaired. Importantly, exercise training restores cAMP-mediated relaxation of collateral-dependent coronary arteries.

Adenosine↗

Dynamics of left ventricular apex rotation during angioplasty: a sensitive index of ischemic dysfunction.

BACKGROUND: Apex rotation has been shown to provide a reliable index of the dynamics of left ventricular (LV) twist. In this study, we aimed to characterize twist at baseline and during acute ischemia in 20 patients undergoing percutaneous transluminal coronary angioplasty to the left anterior descending (LAD) artery and to test whether an old myocardial infarction or collateral flow affected twist dynamics. METHODS AND RESULTS: Among patients with no previous infarction, five had no collaterals (group A) and six had angiographically visible collaterals (group B). Previous anterior infarction was present in nine patients (group C). Data were acquired with the LAD angioplasty wire passed beyond the apex using a view aligned with the LV long axis. Frame-by-frame dynamics of apex rotation were measured from the angular movement of the portion of the wire that traversed the apex. Aortic pressure recordings allowed precise temporal definition of the cardiac cycle. Dynamics of apex rotation were measured at fixed intervals until 60 seconds of occlusion and up to 60 seconds of reperfusion. In group A, counterclockwise apex rotation (twist) during ejection of -22.0+/-1.7 degrees (mean+/-SEE) was followed by rapid clockwise rotation (untwist) during isovolumic relaxation. During 60 seconds of ischemia, maximum apex rotation decreased to -8.2+/-2.0 degrees (P<.001 versus baseline). In group B, baseline apex rotation was similar (-26.2+/-6.9 degrees) to that in group A, but ischemia had less effect, with apex rotation values of -17.7+/-3.4 degrees (P<.05 versus group A values). Group C was characterized by reduced baseline apex rotation values (-9.7+/-3.1 degrees, P<.05 versus group A values), with little change observed during ischemia (-8.1+/-2.6 degrees). CONCLUSIONS: Apex rotation, an index of ventricular twist, is sensitive to acute ischemia in patients without previous myocardial infarction. Visible collaterals to the ischemic region attenuate the acute ischemic response at 60 seconds. Previous myocardial infarction causes abnormalities in the baseline twist pattern with no further deterioration at 60 seconds of ischemia.

Adult↗

Pedestrian injuries: viscoelastic properties of human knee ligaments at high loading rates.

OBJECTIVE: Accidents involving pedestrians are very common, and often lead to severe injuries to the lower extremities. In a large portion of pedestrian-automobile collisions, knee ligament injuries are sustained. In this study, the viscoelastic properties of the four major human knee ligaments were investigated at loading rates representative for pedestrian-automobile collisions. METHODS: Bone-ligament-bone specimens were tested in knee distraction loading. The collateral ligaments and the separate functional bundles of the cruciate ligaments were tested in the anatomical position corresponding to a fully extended knee. A series of step-and-hold tests and ramp tests at different rates were conducted to characterize the time-dependent behavior of the knee ligaments for deformation rates associated with the pedestrian impact loading environment. The quasi linear viscoelastic (QLV) theory was used to describe the structural response of the knee ligaments and averaged parameters for this model were determined. RESULTS: The QLV theory was found to be applicable for the time range that is relevant for pedestrian-automobile collisions. The structural behavior of the knee ligaments was found to be particularly rate-sensitive for high elongation rates, as occur during these collisions. The ligament stiffness was found to increase with age for both the collateral ligaments and with weight for the medial collateral ligament. CONCLUSIONS: For the loading conditions that are relevant for pedestrian-automobile collisions, the use of the QLV model for the description of the mechanical behavior of knee ligaments is appropriate. The rate-sensitivity is particularly important for these extreme loading conditions. The relaxation behavior was found to be consistent between different ligament types and samples. Variations due to donor anthropometry were found predominantly for the instantaneous elastic behavior.

Accidents, Traffic↗

Serotonin blocks different patterns of low Mg2+-induced epileptiform activity in rat entorhinal cortex, but not hippocampus.

Low Mg2+-induced epileptiform activity in the entorhinal cortex is characterized by an initial expression of seizure-like events followed by late recurrent discharges. Both these forms of activity as well as the transition between them were blocked by serotonin. In contrast, serotonin had little effect upon the epileptiform activity in areas CA3 and CA1 of the hippocampus. Both forms of epileptiform activity in the entorhinal cortex are sensitive to N-methyl-D-aspartate receptor antagonists and it is shown here that serotonin blocked both types of epileptiform activity through an effective concentration-dependent reduction of N-methyl-D-aspartate receptor-mediated excitatory postsynaptic potentials in deep layer entorhinal cortex cells. Serotonin also prolonged or even prevented the transition between the two types of epileptiform activity and we suggest that this may be through activation of the Na+/K+-ATPase. The resistance of epileptiform activity in CA1 and CA3 to serotonin was most likely related to the inability of serotonin to reduce Schaffer collateral-evoked excitatory postsynaptic potentials. Given the strong serotonergic inputs to both the hippocampus and entorhinal cortex, the differential sensitivity of the two regions to serotonin suggests functional differences. In addition since the late recurrent discharges in the entorhinal cortex are resistant to all clinically used anticonvulsants, serotonin may open new avenues for the development of novel anticonvulsant compounds.

Animals↗

Retrospective postmortem dementia assessment. Validation of a new clinical interview to assist neuropathologic study.

Neuropathologic studies of dementia and normal aging suffer from a lack of individuals examined for the presence and severity of dementia before death. To increase clinical information in such cases, a retrospective collateral interview was developed. Thirty-nine individuals were studied; 27 had autopsies. In all cases, the autopsy confirmed the Retrospective Collateral Dementia Interview (RCDI) diagnosis of the presence or absence of dementia; the RCDI had a sensitivity of 88% and a specificity of 80% for specifically detecting probable Alzheimer's disease. Agreement between the RCDI and premortem diagnosis was 96%; between RCDI and medical records, 100%. Agreement between RCDI staging of dementia severity and the last assessment of the living subject was 70%; between the RCDI and a brief staging at death, 86%. This validation confirms the value of postmortem interviews with close informants to assess dementia presence and severity.

Aged↗

An evaluation of the quasi-linear viscoelastic properties of the healing medial collateral ligament in a goat model.

The viscoelastic properties of the healing medial collateral ligament (MCL) at 12 weeks after isolated injury were investigated in a goat model. The stress-strain relationships, static and cyclic stress-relaxation behaviors of the healing MCL up to 5% strain were determined experimentally using a femur-MCL-tibia complex. These experimental data were used in combination with the quasi-linear viscoelastic (QLV) theory of Fung (1972) to characterize the reduced relaxation function, G(t) (described by constants C, tau1, and tau2) and the elastic response, sigmae(epsilon) (described by constants A and B) of this tissue. It was found that the percentage of stress relaxation for the healing MCLs was significantly greater than those for sham-operated controls (49.0 +/- 12.1% vs. 26.5 +/- 8.1%, respectively; p < 0.05). The product of constants A x B, i.e. the initial slope of the stress-strain curves, was found to be significantly lower for healing MCLs compared to those for sham-operated controls (32.9 +/- 15.8 MPa vs. 118.8 +/- 48.3 MPa; p < 0.05). The dimensionless constant C, i.e. the magnitude of the viscous response, was nearly three times greater for healing MCLs, while constant tau1 was found to be similar between the two groups (0.80 +/- 0.43 s vs. 0.89 +/- 0.52 s, respectively). Constant tau2 for the healing MCL was significantly less than the controls (1269 +/- 38 s vs. 1845 +/- 431 s; p < 0.05) indicating that the stress relaxation reached a plateau earlier. These constants of the QLV theory used to describe the healing MCL were validated for the strain level utilized in this experiment (approximately equal to 4.5%) by predicting the peak stresses during a cyclic stress-relaxation experiment. The theoretically determined values closely matched the experimentally measured values. Thus, this study demonstrates that the QLV theory could be successfully used to describe the viscoelastic behavior of the MCL during the early phases of healing.

Animals↗

[MRI of knee ligaments: error analysis with reference to meniscus and anterior cruciate ligaments in an arthroscopic controlled patient cohort].

PURPOSE: To categorise discrepancies in findings of the menisci and anterior cruciate ligament (ACL) between arthroscopy and MRI. MATERIALS AND METHODS: The MRIs of 236 patients were retrospectively analysed by an experienced radiologist without knowledge of clinical and/or operative findings. Discrepancies in arthroscopic findings were reevaluated together with the arthroscopist to determine their cause of error. RESULTS: The diagnostic accuracies for injuries of the medial and lateral meniscus and the ACL were 92.4%, 92.4%, and 94.1%, respectively. For the menisci, causes for discrepancies in findings (n = 31) were: overinterpretation of central signal intensities with contact to the meniscal surface but without disturbance of the meniscal contour as a tear (n = 12), insufficient arthroscopie evaluation of the knee joint (n = 11), overlooked tears on MR imaging (n = 6), misinterpretation of normal anatomic structures (n = 1), "magic angle" phenomenon (n = 1), and missed tears at MRI (n = 1). Causes for discrepancies for the ACL (n = 18) were: nearly complete versus complete rupture either at MRI or arthroscopy and vice versa (n = 9), insufficient arthroscopic evaluation (n = 6), insufficient MRI technique (n = 2), and overlooked tear on MR imaging (n = 1). CONCLUSIONS: Discrepant findings between MRI and arthroscopy may be also due to an insufficient arthroscopic evaluation in clinical routine. The close cooperation between surgeons and radiologists improves the understanding of the methods of each other.

Adolescent↗