Müller's Muscles: Three Different Smooth Muscles of the Eye and Orbit

Background: We previously reported that the supratarsal Mueller's muscle is innervated by both sympathetic efferent fibers and trigeminal proprioceptive afferent fibers, which function as mechanoreceptors-inducing reflexive contractions of both the levator and frontalis muscles. Controversy still persists regarding the role of the mechanoreceptors in Mueller's muscle; therefore, we clinically and histologically investigated Mueller's muscle. Methods: We evaluated the role of phenylephrine administration into the upper fornix in contraction of Mueller's smooth muscle fibers and how intraoperative stretching of Mueller's muscle alters the degree of eyelid retraction in 20 patients with aponeurotic blepharoptosis. In addition, we stained Mueller's muscle in 7 cadavers with antibodies against α-smooth muscle actin, S100, tyrosine hydroxylase, c-kit, and connexin 43. Results: Maximal eyelid retraction occurred approximately 3.8 minutes after administration of phenylephrine and prolonged eyelid retraction for at least 20 minutes after administration. Intraoperative stretching of Mueller's muscle increased eyelid retraction due to its reflexive contraction. The tyrosine hydroxylase antibody sparsely stained postganglionic sympathetic nerve fibers, whereas the S100 and c-kit antibodies densely stained the interstitial cells of Cajal (ICCs) among Mueller's smooth muscle fibers. A connexin 43 antibody failed to stain Mueller's muscle. Conclusions: A contractile network of ICCs may mediate neurotransmission within Mueller's multiunit smooth muscle fibers that are sparsely innervated by postganglionic sympathetic fibers. Interstitial cells of Cajal may also serve as mechanoreceptors that reflexively contract Mueller's smooth muscle fibers, forming intimate associations with intramuscular trigeminal proprioceptive fibers to induce reflexive contraction of the levator and frontalis muscles.

Refined distribution of myelinated trigeminal proprioceptive nerve fibres in Mueller's muscle as the mechanoreceptors to induce involuntary reflexive contraction of the levator and frontalis muscles

Neuroanatomical Structures in Extraocular Muscles and Their Potential Implication in the Management of Strabismus

Functional coupling between inositol (1,4,5)-trisphosphate receptor (IP(3)R) and ryanodine receptor (RyR) represents a critical component of intracellular Ca(2+) signaling in many excitable cells; however, the role of this mechanism in skeletal muscle remains elusive. In skeletal muscle, RyR-mediated Ca(2+) sparks are suppressed in resting conditions, whereas application of transient osmotic stress can trigger activation of Ca(2+) sparks that are restricted to the periphery of the fiber. Here we show that onset of these spatially confined Ca(2+) sparks involves interaction between activation of IP(3)R and RyR near the sarcolemmal membrane. Pharmacological prevention of IP(3) production or inhibition of IP(3)R channel activity abolishes stress-induced Ca(2+) sparks in skeletal muscle. Although genetic ablation of the type 2 IP(3)R does not appear to affect Ca(2+) sparks in skeletal muscle, specific silencing of the type 1 IP(3)R leads to ablation of stress-induced Ca(2+) sparks. Our data indicate that membrane-delimited signaling involving cross-talk between IP(3)R1 and RyR1 contributes to Ca(2+) spark activation in skeletal muscle.

The periorbital sheath serves as a major pathway for sympathetic nerves traveling to distal orbital targets in the rat. This tissue accommodates sympathetic fiber sprouting in the neonate but becomes impassable by postnatal day 30 (PND 30). In contrast, smooth muscle target remains receptive to sympathetic ingrowth. To determine the attributes of receptive and nonreceptive tissues, we compared periorbital pathway and target tissue phenotypes prior to (PND 5 and PND 15) and after (PND 30 and PND 60) the period when pathway receptivity is lost. Both pathway cells and superior tarsal smooth muscle cells expressed alpha-smooth muscle actin and smooth muscle myosin heavy chain throughout development. At PND 5-15, both tissues also expressed vimentin, collagen IV, laminin 1 and laminin beta2, whereas fibronectin was detected only in pathway tissue. At PND 30, vimentin, collagen IV, and fibronectin were absent in tarsal muscle but were robust in pathway tissue. Laminin 1 and laminin beta2 expression was maintained in muscle; however, in pathway cells, laminin 1 declined modestly, and laminin beta2 decreased precipitously to barely detectable levels. Quantitative competitive polymerase chain reaction showed that nerve growth factor mRNA was present in the pathway throughout development at levels that were greater than both surrounding connective tissue and tarsal muscle. We conclude that the loss of pathway receptivity to sympathetic nerve ingrowth is associated with a transition from a phenotype similar to fetal smooth muscle cells to one that is more consistent with myofibroblast-like cells.

In an attempt both to preserve and — if possible — to enhance the amplitude of action of Muller's superior tarsal muscle, as a supplement to levator surgery for blepharoptosis, a procedure in which the former muscle, with adherent conjunctiva, is dissected laterally and freed at its origin, prior to levator surgery, to be re-apposed to the back of the latter muscle as high as possible, at the conclusion of the levator procedure, has been tried on five cases, with good preservation of staring and of response to topical epinephrine — to which Muller's muscle but not levator, responds. Preservation of response to topical cocaine as well further implies that the sympathetic innervation of Muller's muscle is also unharmed by the technique, and indicates that its unknown anatomical pathway most probably follows the peripheral arterial arcade of the upper pretarsal space, entering the muscle near its insertion — the only part of Muller's muscle left undissected by this procedure.Ptosis patients with reduced ampl...

Airway smooth muscle: A modulator of airway remodeling in asthma

Sensory nerves impair sympathetic reinnervation and recovery of smooth muscle function

Notch and transforming growth factor-beta (TGFbeta) play pivotal roles during vascular development and the pathogenesis of vascular disease. The interaction of these two pathways is not fully understood. The present study utilized primary human smooth muscle cells (SMC) to examine molecular cross-talk between TGFbeta1 and Notch signaling on contractile gene expression. Activation of Notch signaling using Notch intracellular domain or Jagged1 ligand induced smooth muscle alpha-actin (SM actin), smooth muscle myosin heavy chain, and calponin1, and the expression of Notch downstream effectors hairy-related transcription factors. Similarly, TGFbeta1 treatment of human aortic smooth muscle cells induced SM actin, calponin1, and smooth muscle protein 22-alpha (SM22alpha) in a dose- and time-dependent manner. Hairy-related transcription factor proteins, which antagonize Notch activity, also suppressed the TGFbeta1-induced increase in SMC markers, suggesting a general mechanism of inhibition. We found that Notch and TGFbeta1 cooperatively activate SMC marker transcripts and protein through parallel signaling axes. Although the intracellular domain of Notch4 interacted with phosphoSmad2/3 in SMC, this interaction was not observed with Notch1 or Notch2. However, we found that CBF1 co-immunoprecipitated with phosphoSmad2/3, suggesting a mechanism to link canonical Notch signaling to phosphoSmad activity. Indeed, the combination of Notch activation and TGFbeta1 treatment led to synergistic activation of a TGFbeta-responsive promoter. This increase corresponded to increased levels of phosphoSmad2/3 interaction at Smad consensus binding sites within the SM actin, calponin1, and SM22alpha promoters. Thus, Notch and TGFbeta coordinately induce a molecular and functional contractile phenotype by co-regulation of Smad activity at SMC promoters.

To begin the process of forming neural circuits, new neurons first establish their polarity and extend their axon. Axon extension is guided and regulated by highly coordinated cytoskeletal dynamics. Here we demonstrate that in hippocampal neurons, the actin-binding protein caldesmon accumulates in distal axons, and its N-terminal interaction with myosin II enhances axon extension. In cortical neural progenitor cells, caldesmon knockdown suppresses axon extension and neuronal polarity. These results indicate that caldesmon is an important regulator of axon development.

Platelet-derived growth factor BB (PDGF-BB) has been shown to be an extremely potent negative regulator of smooth muscle cell (SMC) differentiation. Moreover, previous studies have demonstrated that the Kruppel-like transcription factor (KLF) 4 potently represses the expression of multiple SMC genes. However, the mechanisms whereby KLF4 suppresses SMC gene expression are not known, nor is it clear whether KLF4 contributes to PDGF-BB-induced down-regulation of SMC genes. The goals of the present studies were to determine the molecular mechanisms by which KLF4 represses expression of SMC genes and whether it contributes to PDGF-BB-induced suppression of these genes. Results demonstrated that KLF4 markedly repressed both myocardin-induced activation of SMC genes and expression of myocardin. KLF4 was rapidly up-regulated in PDGF-BB-treated, cultured SMC, and a small interfering RNA to KLF4 partially blocked PDGF-BB-induced SMC gene repression. Both PDGF-BB and KLF4 markedly reduced serum response factor binding to CArG containing regions within intact chromatin. Finally, KLF4, which is normally not expressed in differentiated SMC in vivo, was rapidly up-regulated in vivo in response to vascular injury. Taken together, results indicate that KLF4 represses SMC genes by both down-regulating myocardin expression and preventing serum response factor/myocardin from associating with SMC gene promoters, and suggest that KLF4 may be a key effector of PDGF-BB and injury-induced phenotypic switching of SMC.

We have been intrigued to find smooth muscle markers within mouse extraocular muscle (EOM) while studying contractile features of the aqueous humour drainage tissues by immunohistochemistry. Smooth muscle is known to be present in connective tissue fascial pulleys ensheathing EOM1 but not in the EOM per se. The mouse has many available engineered strains, shares many biological similarities with primates, and is widely used to model human biology, including that of EOM.2 Herein, we show that classic smooth muscle proteins of alpha-smooth muscle actin (α-SMA), myosin heavy chain (MHC), caldesmon, and tropomyosin are intimately associated with EOM fibre bundles, a finding that may be relevant to better understanding EOM control. We studied C57BL/6 mice (2-3 months, Charles River, Wilmington, MA) that were housed in a temperature-controlled room with 12h light and dark cycles and fed ad libitum. Animal care and use complied with the Institutional Animal Care and Use Committee as well as the Association for Research in Vision and Ophthalmology guidelines. Anterior (insertion) and posterior (retro-equator) portions of four rectus EOM of C57BL/6 mice (n=5) were carefully isolated, quickly embedded in OCT compound and snap-frozen in liquid nitrogen. Eight μm-thick cryosections were fixed with 4% paraformaldehyde, permeabilised and blocked (0.3% Triton X-100+5% BSA), incubated with primary antibodies (Abcam) to α-SMA, MHC non-muscle, caldesmon, and tropomyosin overnight at 4°C, then secondary antibodies and Alexa 568-phalloidin. Vascular smooth muscle labeling of the same tissues represented positive controls, while normal IgG isotype labeling represented negative controls. Sections were analysed by Leica SP5 or Zeiss LSM 710 confocal microscopy. The same tissue slides were further processed for hematoxylin and eosin staining to confirm structure. Figure 1 shows representative longitudinal and cross sectional immunohistochemistry images of the mouse anterior EOM at their globe insertions. F-actin labeling localized to contractile regions that were mostly consistent with striated muscle fibre bundles. Positive labeling for α-SMA, MHC, and caldesmon labeling was present in the periphery of phalloidin-positive muscle fibre bundles. Positive tropomyosin labeling was seen centrally and peripherally in muscle bundles, corresponding to regions of striated and presumed smooth muscle elements. Smooth muscle marker labeling partially overlapped with phalloidin labeling in fibre bundles. Figure 1 Smooth muscle profile in the anterior part of 4 different rectus extraocular muscles (EOMs). Co-localisation of α-SMA, MHC, caldesmon, or tropomyosin with phalloidin in the anterior portion of EOM. All 4 different rectus EOMs demonstrated a similar ... Figure 2 shows the distribution of smooth muscle markers in representative longitudinal and cross sections of rectus muscles posteriorly. All recti showed this pattern. As in the anterior EOM, α-SMA, MHC, and caldesmon labeling localized to the periphery of phalloidin-labeled bundles, while tropomyosin was present peripherally and centrally. Positive immunolabeling using the same prior antibody panel was seen in vascular smooth muscle from the same tissues (not shown). Figure 2 Smooth muscle profile of the posterior part of 4 different rectus EOMs. Co-localisation of α-SMA, MHC, caldesmon, or tropomyosin with phalloidin in the posterior portion of EOM was shown. All 4 different rectus EOMs demonstrated a similar in situ ... Orbital smooth muscle is present in superior and inferior palpebral muscles, inferior orbital fissure and EOM fascial pulleys1 as part of a periorbital smooth muscle network.5 To our best knowledge smooth muscle has not been described in EOM, which is considered striated muscle. Here we describe classic smooth muscle markers within mouse EOM, mostly in the periphery of striated muscle fiber bundles. Smooth muscle was present in all rectus muscles at their insertions and retro-equatorially, indicating this organization is widely present in mouse EOM. The EOM and their supporting structures represent a complex that helps orchestrate eye movement. Striated EOM are organized as global and orbital layers, each with different structural, vascular, neural, mechanical and metabolic features.3-4 Fascial sheaths and pulleys influence muscle actions on the globe1 under possible smooth muscle modulation. As with autonomically-innervated palpebral smooth muscle that works with striated muscle to regulate eyelid position,5 we postulate that smooth muscle associated with striated EOM plays a role in determining eye position in mice, and possibly in humans.

Levator palpebrae superioris muscle (LPSM) and facial muscles comprise fast-twitch fibers (FTFs) and slow-twitch fibers (STFs) but lack muscle spindles required to contract STFs reflexively. Voluntary contractions and microsaccades of FTFs in LPSM stretch mechanoreceptors in superior tarsal muscle (STM) to induce phasic contractions of STFs in LPSM and frontalis muscle via mesencephalic trigeminal nucleus (MTN). They also induce prolonged contractions of STFs in bilateral frontalis and orbital orbicularis oculi muscles and physiological arousal via MTN and rostral locus coeruleus (LC). We hypothesized that stretching of mechanoreceptors in STM also induces prolonged contractions of STFs in other facial expression muscles (FEMs) via rostral LC. To verify this hypothesis, we reported a case series of abnormal contractions of FEMs due to aponeurosis disinsertion and disordered mechanoreceptor stretching. The first and second cases, which showed unilaterally and bilaterally sensitized mechanoreceptors, respectively, recorded increased prolonged contractions of ipsilateral and bilateral grimacing muscles, respectively. The third and fourth cases with asymmetrically and bilaterally desensitized mechanoreceptors experienced asymmetrically and bilaterally decreased prolonged contractions of grimacing and smiling muscles, respectively. Preoperatively and after surgery was performed to adjust mechanoreceptor stretching and reinsert aponeuroses into tarsi, we evaluated prolonged contractions of grimacing and smiling muscles during primary gazing and facial expression movements. Surgery satisfactorily cured abnormal prolonged contractions of grimacing and smiling muscles. Stretching of mechanoreceptors in STM by microsaccades or voluntary contractions of FTFs in LPSM might activate rostral LC via MTN, which tonically or phasically stimulates FEM motor neurons to reflexively contract their STFs, respectively.

The superior tarsal muscle (STM) is a smooth muscle that originates from the undersurface of the levator palpebrae superioris muscle (LPSM) and inserts onto the superior tarsal plate (STP) of the upper eyelid. We have performed a morphometrical investigation of the STM in 49 adult human cadavers (34 males, 15 females). Histological analysis has shown a transitional area between the skeletal striated muscle (LPSM) and the adjacent smooth muscle (STM). We propose an original morphological classification based upon the attachment of STM to the upper border of the STP. Accordingly, we describe four patterns of STM. Pattern 1 (P1) consists of STM attachment to the central portion of the STP. Pattern 2 consists of both medial (P2M) STM attachment to both the central and medial regions of the STP and lateral (P2L) STM attachment to both the central and lateral regions of the STP. Pattern 3 (P3) consists of STM attachment along the whole extent of the STP. Pattern 3 was the most frequently observed pattern (63.27%) followed by patterns P2M (24.49%), P2L (8.16%) and P1 (4.08%). P3 was the predominant pattern in males (73.52%), while in females, both patterns P2M (46.66%) and P3 (40.00%) were equally prevalent. The analysis of paired specimens revealed a symmetrical arrangement in 72.20% of all cases, with the remaining cases (27.80%) displaying left-right STM asymmetries. To the best of our knowledge, this is the first description of the STM asymmetries in the medical literature. This innovative classification provides anatomical parameters for interpreting morphological variations of the STM with relevant applications in both plastic surgery and ophthalmology.

Induction of Neutrophil Gelatinase-Associated Lipocalin in Vascular Injury via Activation of Nuclear Factor-κB