Identification of vagal afferent nerve endings in the mouse colon and their spatial relationship with enterochromaffin cells.

Understanding how the gut communicates with the brain, via sensory nerves, is of significant interest to medical science. Enteroendocrine cells (EEC) that line the mucosa of the gastrointestinal tract release neurochemicals, including the largest quantity of 5-hydroxytryptamine (5-HT). How the release of substances, like 5-HT, from enterochromaffin (EC) cells activates vagal afferent nerve endings is unresolved. We performed anterograde labelling from nodose ganglia in vivo and identified vagal afferent axons and nerve endings in the mucosa of whole-mount full-length preparations of mouse colon. We then determined the spatial relationship between mucosal-projecting vagal afferent nerve endings and EC cells in situ using 3D imaging. The mean distances between vagal afferent nerve endings in the mucosa, or nearest varicosities along vagal afferent axon branches, and the nearest EC cell were 29.6 ± 19.2 µm (n = 107, N = 6) and 25.7 ± 15.2 µm (n = 119, N = 6), respectively. No vagal afferent endings made close contacts with EC cells. The distances between EC cells and vagal afferent endings are many hundreds of times greater than known distances between pre- and post-synaptic membranes (typically 10-20 nm) that underlie synaptic transmission in vertebrates. The absence of any close physical contacts between 5-HT-containing EC cells and vagal afferent nerve endings in the mucosa leads to the inescapable conclusion that the mechanism by which 5-HT release from ECs in the colonic mucosa occurs in a paracrine fashion, to activate vagal afferents.

The gut-brain axis: spatial relationship between spinal afferent nerves and 5-HT-containing enterochromaffin cells in mucosa of mouse colon.

Cross talk between the gastrointestinal tract and brain is of significant relevance for human health and disease. However, our understanding of how the gut and brain communicate has been limited by a lack of techniques to identify the precise spatial relationship between extrinsic nerve endings and their proximity to specific cell types that line the inner surface of the gastrointestinal tract. We used an in vivo anterograde tracing technique, previously developed in our laboratory, to selectively label single spinal afferent axons and their nerve endings in mouse colonic mucosa. The closest three-dimensional distances between spinal afferent nerve endings and axonal varicosities to enterochromaffin (EC) cells, which contain serotonin (5-hydroxytryptamine; 5-HT), were then measured. The mean distances (± standard deviation) between any varicosity along a spinal afferent axon or its nerve ending, and the nearest EC cell, were 5.7 ± 6.0 µm (median: 3.6 µm) and 26.9 ± 18.6 µm (median: 24.1 µm), respectively. Randomization of the spatial location of EC cells revealed similar results to this actual data. These distances are ∼200-1,000 times greater than those between pre- and postsynaptic membranes (15-25 nm) that underlie synaptic transmission in the vertebrate nervous system. Our findings suggest that colonic 5-HT-containing EC cells release substances to activate centrally projecting spinal afferent nerves likely via diffusion, as such signaling is unlikely to occur with the spatial fidelity of a synapse.NEW & NOTEWORTHY We show an absence of close physical contact between spinal afferent nerves and 5-HT-containing EC cells in mouse colonic mucosa. Similar relative distances were observed between randomized EC cells and spinal afferents compared with actual data. This spatial relationship suggests that substances released from colonic 5-HT-containing EC cells are unlikely to act via synaptic transmission to neighboring spinal afferents that relay sensory information from the gut lumen to the brain.

Control of colonic motility using electrical stimulation to modulate enteric neural activity.

Electrical stimulation of the enteric nervous system (ENS) is an attractive approach to modify gastrointestinal transit. Colonic motor complexes (CMCs) occur with a periodic rhythm, but the ability to elicit a premature CMC depends, at least in part, upon the intrinsic refractory properties of the ENS, which are presently unknown. The objectives of this study were to record myoelectric complexes (MCs, the electrical correlates of CMCs) in the smooth muscle and 1) determine the refractory periods of MCs, 2) inform and evaluate closed-loop stimulation to repetitively evoke MCs, and 3) identify stimulation methods to suppress MC propagation. We dissected the colon from male and female C57BL/6 mice, preserving the integrity of intrinsic circuitry while removing the extrinsic nerves, and measured properties of spontaneous and evoked MCs in vitro. Hexamethonium abolished spontaneous and evoked MCs, confirming the necessary involvement of the ENS for electrically evoked MCs. Electrical stimulation reduced the mean interval between evoked and spontaneous CMCs (24.6 ± 3.5 vs. 70.6 ± 15.7 s, P = 0.0002, n = 7). The absolute refractory period was 4.3 s (95% confidence interval (CI) = 2.8-5.7 s, R2 = 0.7315, n = 8). Electrical stimulation applied during fluid distention-evoked MCs led to an arrest of MC propagation, and following stimulation, MC propagation resumed at an increased velocity (n = 9). The timing parameters of electrical stimulation increased the rate of evoked MCs and the duration of entrainment of MCs, and the refractory period provides insight into timing considerations for designing neuromodulation strategies to treat colonic dysmotility.NEW & NOTEWORTHY Maintained physiological distension of the isolated mouse colon induces rhythmic cyclic myoelectric complexes (MCs). MCs evoked repeatedly by closed-loop electrical stimulation entrain MCs more frequently than spontaneously occurring MCs. Electrical stimulation delivered at the onset of a contraction temporarily suppresses the propagation of MC contractions. Controlled electrical stimulation can either evoke MCs or temporarily delay MCs in the isolated mouse colon, depending on timing relative to ongoing activity.

Identification of a novel distension-evoked motility pattern in the mouse uterus.

The dynamic changes in uterine contractility in response to distension are incompletely understood. Rhythmic, propagating contractions of nonpregnant uterine smooth muscle occur in the absence of nerve activity (i.e., myogenic), events that decline during pregnancy and reemerge at parturition. We therefore sought to determine how myogenic contractions of the nonpregnant uterus are affected by distension, which might provide mechanistic clues underlying distension-associated uterine conditions such as preterm birth. Uteri isolated from nulliparous adult female mice in proestrus were video imaged to generate spatiotemporal maps, and myoelectrical activity simultaneously recorded using extracellular suction electrodes. Motility patterns were examined under basal conditions and following ramped intraluminal distension with fluid to 5 and 10 cmH2O. Intraluminal distension caused pressure-dependent changes in the frequency, amplitude, propagation speed, and directionality of uterine contractions, which reversed upon pressure release. Altered burst durations of underlying smooth muscle myoelectric events were concurrently observed, although action potential spike intervals were unchanged. Voltage-gated sodium channel blockade [tetrodotoxin (TTX); 0.6 µM] attenuated both the amplitude of contractions and burst duration of action potentials, whereas all activity was abolished by L-type calcium channel blockade (nifedipine; 1 µM). These data suggest that myogenic motility patterns of the nonpregnant mouse uterus are sensitive to changes in intraluminal pressure and, at high pressures, may be modulated by voltage-gated sodium channel activity. Future studies may investigate whether similar distension-evoked changes occur in the pregnant uterus and the possible pathophysiological role of such activity in the development of preterm birth.

Diversity of neurogenic smooth muscle electrical rhythmicity in mouse proximal colon.

The mechanisms underlying electrical rhythmicity in smooth muscle of the proximal colon are incompletely understood. Our aim was to identify patterns of electrical rhythmicity in smooth muscle of the proximal region of isolated whole mouse colon and characterize their mechanisms of origin. Two independent extracellular recording electrodes were used to record the patterns of electrical activity in smooth muscle of the proximal region of whole isolated mouse colon. Cross-correlation analysis was used to quantify spatial coordination of these electrical activities over increasing electrode separation distances. Four distinct neurogenic patterns of electrical rhythmicity were identified in smooth muscle of the proximal colon, three of which have not been identified and consisted of bursts of rhythmic action potentials at 1-2 Hz that were abolished by hexamethonium. These neurogenic patterns of electrical rhythmicity in smooth muscle were spatially and temporally synchronized over large separation distances (≥2 mm rosto-caudal axis). Myogenic slow waves could be recorded from the same preparations, but they showed poor spatial and temporal coordination over even short distances (≤1 mm rostro-caudal axis). It is not commonly thought that electrical rhythmicity in gastrointestinal smooth muscle is dependent upon the enteric nervous system. Here, we identified neurogenic patterns of electrical rhythmicity in smooth muscle of the proximal region of isolated mouse colon, which are dependent on synaptic transmission in the enteric nervous system. If the whole colon is studied in vitro, recordings can preserve novel neurogenic patterns of electrical rhythmicity in smooth muscle.NEW & NOTEWORTHY Previously, it has not often been thought that electrical rhythmicity in smooth muscle of the gastrointestinal tract is dependent upon the enteric nervous system. We identified patterns of electrical rhythmicity in smooth muscle of the mouse proximal colon that were abolished by hexamethonium and involved the temporal synchronization of smooth muscle membrane potential over large spatial fields. We reveal different patterns of electrical rhythmicity in colonic smooth muscle that are dependent on the ENS.

Sensory nerve endings arising from single spinal afferent neurons that innervate both circular muscle and myenteric ganglia in mouse colon: colon-brain axis.

There is considerable interest in understanding how contents within the gut wall (including microbiome) can activate sensory nerve endings in the gut that project to the central nervous system. However, we have only recently begun to understand the location and characteristics of extrinsic spinal afferent nerve endings that innervate the lower gastrointestinal (GI) tract. Our aim is to identify the nerve endings in the mouse distal colon that arise from single spinal afferent neurons. C57BL/6 mice were anaesthetised and single dorsal root ganglia (DRG) between lumbosacral L6-S1 were injected with dextran biotin. Mice recovered for 7 days. Animals were then euthanized and whole colons removed, fixed and stained for calcitonin-gene-related-peptide (CGRP). Single spinal afferent nerve axons were identified entering the distal colon that ramified along many rows of myenteric ganglia, often giving rise to varicose nerve endings. These same axons bifurcated in the circular muscle giving rise to 4-5 groups of branching-type intramuscular endings, where each group of endings was separated by ~ 370 µm in the rostro-caudal axis and projected 1.2 mm around the circumference. As spinal afferent axons bifurcated, their axons often showed dramatic reductions in diameter. Here, we identified in the distal colon, the characteristics of nerve endings that arise from single colorectal-projecting axons with cell bodies in DRG. These findings suggest that a population of sensory neurons in DRG can potentially detect sensory stimuli simultaneously via different morphological types of endings that lie in both colonic smooth muscle and myenteric ganglia.

Identification of a Rhythmic Firing Pattern in the Enteric Nervous System That Generates Rhythmic Electrical Activity in Smooth Muscle.

The enteric nervous system (ENS) contains millions of neurons essential for organization of motor behavior of the intestine. It is well established that the large intestine requires ENS activity to drive propulsive motor behaviors. However, the firing pattern of the ENS underlying propagating neurogenic contractions of the large intestine remains unknown. To identify this, we used high-resolution neuronal imaging with electrophysiology from neighboring smooth muscle. Myoelectric activity underlying propagating neurogenic contractions along murine large intestine [also referred to as colonic migrating motor complexes, (CMMCs)] consisted of prolonged bursts of rhythmic depolarizations at a frequency of ∼2 Hz. Temporal coordination of this activity in the smooth muscle over large spatial fields (∼7 mm, longitudinally) was dependent on the ENS. During quiescent periods between neurogenic contractions, recordings from large populations of enteric neurons, in mice of either sex, revealed ongoing activity. The onset of neurogenic contractions was characterized by the emergence of temporally synchronized activity across large populations of excitatory and inhibitory neurons. This neuronal firing pattern was rhythmic and temporally synchronized across large numbers of ganglia at ∼2 Hz. ENS activation preceded smooth muscle depolarization, indicating rhythmic depolarizations in smooth muscle were controlled by firing of enteric neurons. The cyclical emergence of temporally coordinated firing of large populations of enteric neurons represents a unique neural motor pattern outside the CNS. This is the first direct observation of rhythmic firing in the ENS underlying rhythmic electrical depolarizations in smooth muscle. The pattern of neuronal activity we identified underlies the generation of CMMCs.SIGNIFICANCE STATEMENT How the enteric nervous system (ENS) generates neurogenic contractions of smooth muscle in the gastrointestinal (GI) tract has been a long-standing mystery in vertebrates. It is well known that myogenic pacemaker cells exist in the GI tract [called interstitial cells of Cajal (ICCs)] that generate rhythmic myogenic contractions. However, the mechanisms underlying the generation of rhythmic neurogenic contractions of smooth muscle in the GI tract remains unknown. We developed a high-resolution neuronal imaging method with electrophysiology to address this issue. This technique revealed a novel pattern of rhythmic coordinated neuronal firing in the ENS that has never been identified. Rhythmic neuronal firing in the ENS was found to generate rhythmic neurogenic depolarizations in smooth muscle that underlie contraction of the GI tract.

Synaptic activation of putative sensory neurons by hexamethonium-sensitive nerve pathways in mouse colon.

The gastrointestinal tract contains its own independent population of sensory neurons within the gut wall. These sensory neurons have been referred to as intrinsic primary afferent neurons (IPANs) and can be identified by immunoreactivity to calcitonin gene-related peptide (CGRP) in mice. A common feature of IPANs is a paucity of fast synaptic inputs observed during sharp microelectrode recordings. Whether this is observed using different recording techniques is of particular interest for understanding the physiology of these neurons and neural circuit modeling. Here, we imaged spontaneous and evoked activation of myenteric neurons in isolated whole preparations of mouse colon and correlated recordings with CGRP and nitric oxide synthase (NOS) immunoreactivity, post hoc. Calcium indicator fluo 4 was used for this purpose. Calcium responses were recorded in nerve cell bodies located 5-10 mm oral to transmural electrical nerve stimuli. A total of 618 recorded neurons were classified for CGRP or NOS immunoreactivity. Aboral electrical stimulation evoked short-latency calcium transients in the majority of myenteric neurons, including ~90% of CGRP-immunoreactive Dogiel type II neurons. Activation of Dogiel type II neurons had a time course consistent with fast synaptic transmission and was always abolished by hexamethonium (300 µM) and by low-calcium Krebs solution. The nicotinic receptor agonist 1,1-dimethyl-4-phenylpiperazinium iodide (during synaptic blockade) directly activated Dogiel type II neurons. The present study suggests that murine colonic Dogiel type II neurons receive prominent fast excitatory synaptic inputs from hexamethonium-sensitive neural pathways. NEW & NOTEWORTHY Myenteric neurons in isolated mouse colon were recorded using calcium imaging and then neurochemically defined. Short-latency calcium transients were detected in >90% of calcitonin gene-related peptide-immunoreactive neurons to electrical stimulation of hexamethonium-sensitive pathways. Putative sensory Dogiel type II calcitonin gene-related peptide-immunoreactive myenteric neurons may receive widespread fast synaptic inputs in mouse colon.

A Contact Pressure Analysis Comparing an All-Inside and Inside-Out Surgical Repair Technique for Bucket-Handle Medial Meniscus Tears.

PURPOSE: To directly compare effectiveness of the inside-out and all-inside medial meniscal repair techniques in restoring native contact area and contact pressure across the medial tibial plateau at multiple knee flexion angles. METHODS: Twelve male, nonpaired (n = 12), fresh-frozen human cadaveric knees underwent a series of 5 consecutive states: (1) intact medial meniscus, (2) MCL tear and repair, (3) simulated bucket-handle longitudinal tear of the medial meniscus, (4) inside-out meniscal repair, and (5) all-inside meniscal repair. Knees were loaded with a 1,000-N axial compressive force at 5 knee flexion angles (0°, 30°, 45°, 60°, 90°), and contact area, mean contact pressure, and peak contact pressure were calculated using thin film pressure sensors. RESULTS: No significant differences were observed between the inside-out and all-inside repair techniques at any flexion angle for contact area, mean contact pressure, and peak contact pressure (all P > .791). Compared with the torn meniscus state, inside-out and all-inside repair techniques resulted in increased contact area at all flexion angles (all P < .005 and all P < .037, respectively), decreased mean contact pressure at all flexion angles (all P < .007 and all P < .001, respectively) except for 0° (P = .097 and P = .39, respectively), and decreased peak contact pressure at all flexion angles (all P < .001, all P < .001, respectively) except for 0° (P = .080 and P = .544, respectively). However, there were significant differences in contact area and peak contact pressure between the intact state and inside-out technique at angles ≥45° (all P < .014 and all P < .032, respectively). Additionally, there were significant differences between the intact state and all-inside technique in contact area at 60° and 90° and peak contact pressure at 90° (both P < .005 and P = .004, respectively). Median values of intact contact area, mean contact pressure, and peak contact pressure over the tested flexion angles ranged from 498 to 561 mm2, 786 to 997 N/mm2, and 1,990 to 2,215 N/mm2, respectively. CONCLUSIONS: Contact area, mean contact pressure, and peak contact pressure were not significantly different between the all-inside and inside-out repair techniques at any tested flexion angle. Both techniques adequately restored native meniscus biomechanics near an intact level. CLINICAL RELEVANCE: An all-inside repair technique provided similar, native-state-restoring contact mechanics compared with an inside-out repair technique for the treatment of displaced bucket-handle tears of the medial meniscus. Thus, both techniques may adequately decrease the likelihood of cartilage degeneration.

Quantitative and Computed Tomography Anatomic Analysis of Glenoid Fixation for Superior Capsule Reconstruction: A Cadaveric Study.

PURPOSE: To investigate glenoid fixation for superior capsule reconstruction (SCR) and evaluate anchor positions, intraosseous trajectories, and proximity to the suprascapular nerve (SSN) and glenoid fossa. The secondary purpose was to provide technical pearls and pitfalls for anchor insertion on the superior glenoid during SCR. METHODS: Three beath pins were arthroscopically inserted into 12 (n = 12) nonpaired human cadaveric shoulders through Neviaser, anterior, and posterior portals to simulate anchor placement on the superior glenoid during SCR. Computed tomography scans were performed to evaluate anchor positioning and insertion trajectories. Specimens were then dissected to delineate the anatomic relations of the beath pins to the SSN and glenoid fossa. RESULTS: The superior glenoid anchor position was a mean 15.0 ± 4.0 mm to the SSN and 6.5 ± 1.7 mm to the glenoid fossa. The posterior glenoid anchor position was a mean 11.8 ± 2.1 mm to the SSN and 2.9 ± 2.9 mm to the glenoid fossa. On average, the superior pin was placed at 12:30 ± 0:30 (left-sided glenoid clock face) and inserted at 19° ± 9° with respect to the sagittal plane of the glenoid, the anterior pin was placed at 11:00 ± 0:30 and inserted 40° ± 17° off the glenoid, and the posterior pin was placed at 3:00 ± 1:00 and inserted at 52° ± 12° off the glenoid. CONCLUSIONS: The results of the present cadaveric study showed that glenoid fixation was safe with respect to the SSN and delineated technical guidelines and trajectories for inserting 3 anchors into the glenoid. CLINICAL RELEVANCE: This study shows that 3 anchors can be inserted into the glenoid without a risk of SSN damage and delineates technical guidelines for anchor insertion.

Comparison of Radiographs and Computed Tomography for the Screening of Anterior Inferior Iliac Spine Impingement.

PURPOSE: To compare radiographic and 3-dimensional (3D) computed tomography (CT) imaging modalities for the screening of anterior inferior iliac spine (AIIS) impingement by establishing imaging measurement related to the AIIS. METHODS: Anteroposterior and false-profile radiographs and 3D CT scans were obtained on 10 human cadaveric pelvises. On the anteroposterior view for each methodology, 2 measurements were calculated: distance to the most lateral AIIS from the 12 o'clock position on the acetabular rim, and the angle between the lateral AIIS and the sagittal plane. On the false-profile view for each methodology, 2 measurements were calculated: distance to the anterior AIIS from the 12 o'clock position on the acetabular rim, and the angle between the anterior AIIS and the sagittal plane. Inter-rater and intrarater reliability analyses were performed for both methods in addition to an intermethod analysis. RESULTS: The radiographic false-profile view was the most repeatable orientation, with intraclass correlation coefficients showing excellent reproducibility in both inter-rater (angle: 0.980, distance: 0.883) and intrarater (angle: 0.995, distance: 0.995) analyses. The mean distance from the 12 o'clock position of the acetabular rim to the most anterior/lateral aspect of the AIIS was 41.4 mm and 16.0 mm on the radiographic false-profile and anteroposterior views, respectively. Intermethod analysis showed a systematic, quantitative bias between modalities (anteroposterior view: -4.1 mm, 6.7°; false-profile view: -0.1 mm, 8.3°), which will remain relatively consistent as evidenced by the strong individual reproducibility of each measurement. CONCLUSIONS: AIIS morphology in relation to the acetabular rim 12 o'clock position and its angle relative to the sagittal plane can be quantitatively determined using either radiographic or 3D CT imaging modalities. CLINICAL RELEVANCE: Radiographic evaluation may be a valuable tool in the screening of AIIS impingement.

Impact of Arthroscopic Lateral Acromioplasty on the Mechanical and Structural Integrity of the Lateral Deltoid Origin: A Cadaveric Study.

PURPOSE: To determine whether a 5-mm and/or 10-mm arthroscopic lateral acromioplasty (ALA) would weaken the structural and mechanical integrity of the lateral deltoid. METHODS: The acromion and lateral deltoid origin were harvested from 15 pairs (n = 30) of fresh-frozen human cadaveric shoulder specimens. One side of each specimen pair (left or right) was randomly assigned to either a 5-mm (n = 7) or 10-mm (n = 8) ALA group, and the contralateral sides (n = 15) were used as matched controls. Acromion thickness and width were measured pre- and postoperatively. After ALA, specimens were inspected for damage to the lateral deltoid origin. Each specimen was secured within a dynamic testing machine, and the deltoid muscle was pulled to failure. Statistical analysis was performed to determine whether ALA reduced the lateral deltoid's failure load. RESULTS: There was no significant difference in failure load between the 5-mm ALA group (661 ± 207 N) and its matched control group (744 ± 212 N; mean difference = 83 N; 95% confidence interval [CI], -91 to 258; P = .285) nor between the 10-mm ALA group (544 ± 210 N) and its matched control group (598 ± 157 N; mean difference = 54 N; 95% CI, -141 to 250; P = .532). There was no correlation found between the amount of bone resected (measured by percent thickness and width of the acromion after ALA) and the failure load of the deltoid. Visual evaluation of the acromion after ALA revealed the lateral deltoid origin had no damage in any case. CONCLUSIONS: ALA did not weaken the structural or mechanical integrity of the lateral deltoid origin. Neither a 5-mm nor a 10-mm ALA significantly reduced the deltoid's failure load. The lateral deltoid origin was not macroscopically damaged in any case. CLINICAL RELEVANCE: ALA can be performed without the potential risk of macroscopically damaging the lateral deltoid origin or reducing its failure load.

Double-bundle posterior cruciate ligament reconstruction: a biomechanical analysis of simulated early motion and partial and full weightbearing on common reconstruction grafts.

PURPOSE: The purpose of this study was to determine the biomechanical effects of simulated immediate motion and weightbearing during rehabilitation on different double-bundle posterior cruciate ligament reconstruction (DB-PCLR) graft options. METHODS: Nine each of commercially prepared (allograft) Achilles tendon allografts, fresh-frozen (autograft) bone-patellar tendon-bone grafts, and fresh-frozen quadriceps tendon grafts were paired with commercially prepared anterior tibialis allografts, fresh-frozen semitendinosus grafts, and fresh-frozen semitendinosus grafts, respectively. Graft pairs were loaded to simulate early range of motion on a stationary bicycle, partial weightbearing (30 %), and full weightbearing. RESULTS: Acquired laxity (displacement, mm) between graft pairs was not significantly different during simulated early range of motion. However, during simulated partial weightbearing, the median acquired laxity of the patellar tendon/semitendinosus pair (1.06 mm) was significantly less than that of the quadriceps tendon/semitendinosus (1.50 mm, p = 0.01) and Achilles/anterior tibialis (1.44 mm, p = 0.003) graft pairs. During simulated full weightbearing, significantly less acquired laxity was observed for the patellar tendon/semitendinosus graft pair (2.38 mm) compared to the Achilles/anterior tibialis pair (4.85 mm, p = 0.04), but a significant difference was not observed compared to the QT/semitendinosus graft pair (3.91 mm, n.s.). There were no significant differences in the ultimate loads between any of the graft pairs. CONCLUSIONS: Simulated early range of motion and early partial weightbearing did not result in clinically significant acquired graft laxity in common graft options utilized for DB-PCLR. However, simulated full weightbearing did result in clinically significant acquired graft laxity, and therefore, early rehabilitation protocols should avoid implementing full weightbearing that could contribute to graft failure.

High-load preconditioning of human soft tissue hamstring grafts: An in vitro biomechanical analysis.

PURPOSE: In order to minimize viscoelastic elongation of ACL reconstruction grafts, preconditioning protocols have been employed in clinical practice prior to final graft fixation. The purpose of this study was to evaluate two separate high-load static preconditioning protocols of double-looped semitendinosus-gracilis grafts and compare these results to both a current clinical protocol and a control group with no preconditioning protocol applied. It was hypothesized that a high-load, static preconditioning protocol would minimize graft elongation during a simulated progressive early rehabilitation compared to both the "89 N" clinical protocol and control groups. METHODS: Grafts were randomly allocated into four preconditioning study groups: (1) control (no preconditioning), (2) clinical protocol (89 N for 15 min), (3) high-load, short duration (600 N for 20 s), and (4) high-load, long duration (600 N for 15 min). After preconditioning, grafts were cyclically loaded between 10 and 400 N at 0.5 Hz for 450 cycles to simulate early postoperative rehabilitation. Graft displacement (elongation) was recorded during both preconditioning and cyclic loading. RESULTS: Increased preconditioning load magnitude and duration significantly reduced graft elongation during cyclic loading (p < 0.05) which corresponded to an inverse relationship with increased elongation during preconditioning. The "600 N for 15 min" protocol resulted in significantly less elongation during simulated early rehabilitation than both the control group and the "89 N for 15 min" protocol (p < 0.001, p < 0.05). CONCLUSIONS: Graft elongation during simulated early rehabilitation was significantly reduced by a high-load preconditioning protocol applied for an extended period of time compared to a current common clinical protocol and grafts that were not preconditioned. In addition, the amount of elongation during simulated early rehabilitation was similar between grafts preconditioned using the current clinical practice protocol and the high-load/short-duration protocol, implying that the latter could potentially induce the same viscoelastic changes in soft tissue grafts as the current clinical practice. The "600 N for 20 s" preconditioning protocol may provide similar postoperative results as the clinical protocol, "89 N for 15 min", and also reduce or maintain operative time. A high-load preconditioning protocol that reduces graft elongation may benefit patients undergoing ACL reconstruction, especially for cases of failed primary reconstruction, genu recurvatum, and increased tibial slope, where maintaining graft length is imperative to restore knee stability.

Computed tomography-based prediction of the straight antegrade humeral nail's entry point and exposure of "critical types": truth or fiction?

BACKGROUND: Straight antegrade intramedullary nailing of proximal humerus fractures has shown promising clinical results. However, up to 36% of all humeri seem to be "critical types" in terms of the potential violation of the supraspinatus (SSP) tendon footprint by the nail's insertion zone. The aims of this study were to evaluate if a computed tomography (CT) scan could reliably predict the nail's entry point on the humeral head and if it would be possible to preoperatively estimate the individual risk of iatrogenic violation of the SSP tendon footprint by evaluating the uninjured contralateral humerus. METHODS: Twenty matched pairs of human cadaveric shoulders underwent CT scans, and the entry point for an antegrade nail as well as measurements regarding critical distances between the entry point and the rotator cuff were determined. Next, gross anatomic measurements of the same data were performed and compared. Furthermore, specimens were reviewed for critical types. RESULTS: Overall, 42.5% of all specimens were found to be critical types. The CT measurements exhibited excellent intra-rater and inter-rater reliability (intraclass correlation coefficients >0.90). Similarly, excellent agreement between the CT scan and gross anatomic measurements in contralateral shoulders (intraclass correlation coefficients >0.88) was found. CONCLUSION: Assessing the uninjured contralateral side, CT can reliably predict the entry point in antegrade humeral nailing and preoperatively identify critical types of humeral heads at risk of iatrogenic implantation damage to the SSP tendon footprint. This study may help surgeons in the decision-making processon which surgical technique should be used without putting the patient at risk for iatrogenic, implant-related damage to the rotator cuff.

Biomechanical evaluation of straight antegrade nailing in proximal humeral fractures: the rationale of the "proximal anchoring point".

PURPOSE: Varus failure is one of the most common failure modes following surgical treatment of proximal humeral fractures. Straight antegrade nails (SAN) theoretically provide increased stability by anchoring to the densest zone of the proximal humerus (subchondral zone) with the end of the nail. The aim of this study was to biomechanically investigate the characteristics of this "proximal anchoring point" (PAP). We hypothesized that the PAP would improve stability compared to the same construct without the PAP. METHODS: Straight antegrade humeral nailing was performed in 20 matched pairs of human cadaveric humeri for a simulated unstable two-part fracture. RESULTS: Biomechanical testing, with stepwise increasing cyclic axial loading (50-N increments each 100 cycles) at an angle of 20° abduction revealed significantly higher median loads to failure for SAN constructs with the PAP (median, 450 N; range, 200-1.000 N) compared to those without the PAP (median, 325 N; range, 100-500 N; p = 0.009). SAN constructs with press-fit proximal extensions (endcaps) showed similar median loads to failure (median, 400 N; range, 200-650 N), when compared to the undersized, commercially available SAN endcaps (median, 450 N; range, 200-600 N; p = 0.240). CONCLUSIONS: The PAP provided significantly increased stability in SAN constructs compared to the same setup without this additional proximal anchoring point. Varus-displacing forces to the humeral head were superiorly reduced in this setting. This study provides biomechanical evidence for the "proximal anchoring point's" rationale. Straight antegrade humeral nailing may be beneficial for patients undergoing surgical treatment for unstable proximal humeral fractures to decrease secondary varus displacement and thus potentially reduce revision rates.

Imaging activation of peptidergic spinal afferent varicosities within visceral organs using novel CGRPα-mCherry reporter mice.

In vertebrates, visceral pain from internal organs is detected by spinal afferents, whose cell bodies lie in dorsal root ganglia (DRG). Until now, all recordings from spinal afferents have been restricted to recording transmission of action potentials along axons, or from cell bodies lying outside their target organ, which is not where sensory transduction occurs. Our aim was to record directly from a major class of spinal afferent within visceral organs, where transduction of sensory stimuli into action potentials occurs. Using novel calcitonin gene-related peptide (CGRP)α reporter mice, DRG neurons expressed mCherry, including nerve axons within viscera. In colon, a minority of total CGRP immunoreactivity was attributed CGRPα. In isolated unstretched colon, calcium imaging from CGRPα-expressing varicose axons did not detect resolvable calcium transients. However, noxious levels of maintained circumferential stretch to the colon induced repetitive calcium transients simultaneously in multiple neighboring varicosities along single mCherry-expressing axons. Discrete varicosities could generate unitary calcium transients independently of neighboring varicosities. However, axons expressing mCherry only generated coordinated calcium transients when accompanied by simultaneous activation of multiple varicosities along that axon. Simultaneous imaging from different classes of myenteric neurons at the same time as mCherry-expressing axons revealed coordinated calcium transients in multiple myenteric neurons, independent of activity in mCherry-expressing axons. CGRPα-expressing axon terminals preferentially responded to heat, capsaicin, and low pH. We show that direct recordings can be made from the major class of peptidergic spinal afferent that contributes to visceral nociception. This approach can provide powerful insights into transduction of stimuli in viscera.

The Effects of Arthroscopic Lateral Acromioplasty on the Critical Shoulder Angle and the Anterolateral Deltoid Origin: An Anatomic Cadaveric Study.

PURPOSE: To investigate if (1) an anterolateral acromioplasty and (2) a lateral acromion resection alter the critical shoulder angle (CSA) without affecting the deltoid origin. METHODS: First, the native CSAs of 10 human cadaveric shoulders (6 male and 4 female specimens; mean age, 54.2 years) were determined with the use of fluoroscopy. Setup allowed for consistent repetitive measurements. Next, a standard arthroscopic anterolateral acromioplasty was performed to create a type 1 acromion, and the CSA was reassessed fluoroscopically. Afterward, a lateral acromioplasty was performed with a 5-mm lateral acromion resection using a 5-mm burr, and the CSA was measured again. The native CSA was compared with (1) the CSA after acromioplasty and (2) the CSA after acromioplasty and lateral acromion resection using a paired t test. Finally, the acromial deltoid attachment was evaluated anatomically for damage to the anterolateral origin. RESULTS: The mean native CSA (34.3° ± 2.1°) was reduced significantly by acromioplasty (33.1° ± 2.0°, P < .001) and further reduced by lateral acromion resection (31.5° ± 1.7°, P < .001). Anterolateral acromioplasty reduced the CSA by a mean of 1.4° (95% confidence interval boundaries, 0.8° and 1.9°), and in combination with lateral acromion resection, the CSA was reduced by a mean of 2.8° (95% confidence interval boundaries, 2.1° and 3.5°). In all specimens (5 of 5) with a presurgery CSA of 35° or greater, the CSA was reduced to the range of 30° to 35° by the combination of both techniques. However, in 2 specimens with a CSA of approximately 32°, the CSA was reduced to less than 30°. The acromial deltoid attachment was found to be well preserved in all specimens. CONCLUSIONS: Arthroscopic anterolateral acromioplasty and a 5-mm lateral acromion resection each reduced the CSA significantly and did not damage the deltoid origin. CLINICAL RELEVANCE: The combination of both techniques could potentially be used in clinical practice to reduce a CSA greater than 35° to the desired range of 30° to 35°.

Biomechanical Results of Lateral Extra-articular Tenodesis Procedures of the Knee: A Systematic Review.

PURPOSE: To systematically review and compare biomechanical results of lateral extra-articular tenodesis (LET) procedures. METHODS: A systematic review was performed using the PubMed, Medline, Embase, and Cochrane databases. The search terms included the following: extraarticular, anterolateral, iliotibial, tenodesis, plasty, augmentation, procedure, reconstruction, technique, biomechanics, kinematic, robot, cadaver, knee, lateral tenodesis, ACL, Marcacci, Lemaire, Losee, Macintosh, Ellison, Andrews, Hughston, and Muller. The inclusion criteria were nonanatomic, in vitro biomechanical studies, defined as in vitro investigations of joint motion resulting from controlled, applied forces. RESULTS: Of the 10 included studies, 7 analyzed anterior tibial translation and reported that isolated LET procedures did not restore normal anterior stability to the anterior cruciate ligament (ACL)-deficient knee. Seven of the 8 studies analyzing tibial rotation reported a reduction in internal tibial rotation across various flexion angles in the ACL-deficient knee when compared with the native state. Five studies reported a reduction in intra-articular graft force with the addition of an LET. Two studies evaluated length change patterns, graft course, and total strain range and found that reconstruction techniques in which the graft attached proximal to the lateral epicondyle and coursed deep to the fibular collateral ligament were most isometric. CONCLUSIONS: In the ACL-deficient knee, LET procedures overconstrained the knee and restricted internal tibial rotation when compared with the native state. In addition, isolated LET procedures did not return normal anterior stability to the ACL-deficient knee but did significantly reduce anterior tibial translation and intra-articular graft forces during anteriorly directed loading. CLINICAL RELEVANCE: Combined injury to the ACL and anterolateral structures has been reported to exhibit greater anterolateral rotatory instability when compared with isolated ACL injuries. Despite the reported risk of joint over-constraint, consideration should be given to reconstructing the anterolateral structures and the ACL concurrently to maximally restore both anterior tibial translation and rotatory stability.

Influence of lateral meniscal posterior root avulsions and the meniscofemoral ligaments on tibiofemoral contact mechanics.

PURPOSE: The purpose of this study was to investigate the effect of lateral meniscal posterior root avulsions combined with intact meniscofemoral ligaments (MFLs), deficient MFLs, anterior cruciate ligament (ACL) tears and reconstructions, and root repairs using an established tibiofemoral contact mechanics testing protocol. METHODS: Ten fresh-frozen cadaveric knees were tested with six knee conditions (1: intact; 2: lateral meniscal posterior root avulsion; 3: root avulsion and deficient MFLs; 4: condition 3 with ACL tear; 5: condition 4 with ACL reconstruction; 6: ACL reconstruction with root repair) at five flexion angles (0°, 30°, 45°, 60°, and 90°), under a 1000-N axial load. Contact area and pressure were measured with Tekscan sensors. RESULTS: Compared to the intact state, condition 2 did not significantly change lateral compartment contact area or pressure. Changes in contact mechanics were greater at increased flexion angles; for condition 3 at 0° and 90°, contact area decreased 37 and 52 % [95 % CI (21-53) and (39-66), respectively] and mean contact pressure increased 55 and 87 % [95 % CI (33-76) and (59-114), respectively]. Root repair with ACL reconstruction was not significantly different from the intact state. CONCLUSIONS: The MFLs protect the lateral compartment from changes in contact mechanics in the setting of a lateral meniscal posterior root avulsion, whereas a combination of lateral meniscal root avulsion and deficient MFLs leads to significant changes. Concurrent ACL reconstruction and lateral meniscal root repair restore mean contact pressure and area to the intact state and are recommended in this combined injury to prevent or slow the development of lateral compartment arthritis.