Gamma-tubulin during the differentiation of spermatozoa in various mammals and man

The distribution of gamma-tubulin as a marker of microtubule organizing centres (MTOC) was studied during spermiogenesis in rodents and in rabbit, monkey and man. A polyclonal antibody directed against human gamma-tubulin was used both for indirect immunofluorescence (IIF) and post-embedding immunogold procedures. In all species, gamma-tubulin was detected in the proximal and distal centrioles of round spermatids. In elongating spermatids, gamma-tubulin was predominantly found in the pericentriolar material (PCM) of both centrioles and particularly around the adjunct of the proximal centriole. At this level, some labelling was also associated with manchette microtubules, but other parts of the manchette and the nuclear ring were never labelled. We propose a role for distal centriole gamma-tubulin in axoneme nucleation and centriolar adjunct gamma-tubulin in manchette nucleation. The disappearance of gamma-tubulin in mature spermatozoa indicates that sperm aster nucleation should be dependent on oocyte gamma-tubulin. Remnants of gamma-tubulin in some human spermatozoa suggest that paternal gamma-tubulin also could contribute to sperm aster formation.     

Behavior of centrioles during meiosis in the male silkworm, Bombyx mori (Lepidoptera)

The behavior of centrioles during eupyrene and apyrene meiosis was examined in the silkworm, Bombyx mori, by transmission electron microscopy and indirect immunofluorescence for tubulin. In eupyrene spermatocytes the centrioles, accompanied by axonemes, attached temporarily to the nucleus at diplotene, then detached from the nucleus in diakinesis. After the separation, a beret-shaped structure consisting of a double membrane covered the proximal region of the pair of centrioles. The structure disappeared after breakdown of the nuclear membrane. The centriole, with the axoneme, reattached to the nucleus at telophase I. The process was repeated during meiosis II until the centrioles maintained their nuclear attachment in newly developed spermatids. In stark contrast to their eupyrene counterparts, apyrene spermatocytes were conspicuously devoid of any attachment of the centrioles to the nucleus. These eupyrene-specific and apyrene-specific relationships were consistently and repeatedly found between the nuclear membrane and centrioles, giving rise to suspicion that the behavioral phenomena may be related to differentiation of the dimorphic sperm types.     

Ultrastructural changes in the cell center during enterocyte differentiation in the mouse

Centrioles in the enterocytes of murine small intestine are located far away from the nuclei and close to the apical surface of cells (1-3 microns from the brush border). Centrioles never form a primary cilium and are not attached to the plasma membrane. Centrosomes (cell center) in enterocytes undergo subsequent involution in respect to the position of cell in the crypt-villus system. Five zones were studied separately: the bottom of the crypt (approx. 1/3 of the appendix), the upper part of the crypt (before its transition into the villus), the basal part, the middle and top of the villus. A centrosome in the crypt contains two centrioles (maternal and daughter) located close to each other. In 10 of 27 cells studied centrioles were replicating. A maternal centriole has 2-3 pericentriolar satellites and appendages. Several cytoplasmic microtubules run towards mother centrioles. Centrioles at the upper part of the crypt split at a distance up to 1.5 microns from each other. The average number of cytoplasmic microtubules and pericentriolar satellites decreases. At the basal part of the villus, centrioles split a distance up to 5 microns from each other; seldom microtubules may be found in a radius of 1 micron around centrioles and none of them is attached to centrioles or satellites. Starting from the middle of the villus, centrioles sometimes may have incomplete triplets of microtubules at the distal and (or) the proximal end. At the upper part of the villus there is only one centriole per cell, that was found in 10 of 50 cells studied on a complete series of 0.2 micron thick sections.     

Ultrastructure of segmentation mitoses in cleaving newt eggs after blocking of centrospheres by quinoline

In quinoline treated blastomeres of Triturus helveticus Raz. and Pleurodeles waltlii Michah., microtubules disappear and around the centrioles markedly enlarged dense bodies accompanied by striated bodies are accumulated, from prophase beyond metaphase. It is concluded that these bodies in untreated cells play a role in the formation of the spindle microtubules. - During metaphase the chromosome are smooth-surfaced and show a pronounced tendency to stickiness. During ana-telophase they become surrounded by nuclear membranes and form caryomeres. - Qinoline does not interfere with centriole replication, but prevents the separation if diplosomes that have been already formed, if it acts before that separation. - Some centrioles exhibit different degrees of ultrastructural disarrangement.     

A molecular marker for centriole maturation in the mammalian cell cycle

The centriole pair in animals shows duplication and structural maturation at specific cell cycle points. In G1, a cell has two centrioles. One of the centrioles is mature and was generated at least two cell cycles ago. The other centriole was produced in the previous cell cycle and is immature. Both centrioles then nucleate one procentriole each which subsequently elongate to full-length centrioles, usually in S or G2 phase. However, the point in the cell cycle at which maturation of the immature centriole occurs is open to question. Furthermore, the molecular events underlying this process are entirely unknown. Here, using monoclonal and polyclonal antibody approaches, we describe for the first time a molecular marker which localizes exclusively to one centriole of the centriolar pair and provides biochemical evidence that the two centrioles are different. Moreover, this 96-kD protein, which we name Cenexin (derived from the Latin, senex for "old man," and Cenexin for centriole) defines very precisely the mature centriole of a pair and is acquired by the immature centriole at the G2/M transition in prophase. Thus the acquisition of Cenexin marks the functional maturation of the centriole and may indicate a change in centriolar potential such as its ability to act as a basal body for axoneme development or as a congregating site for microtubule-organizing material.     

The ultrastructure of the cell center in the enterocytes of mouse embryos and newborn mice

The centrosome structure was studied in differentiating small intestine enterocytes of mouse embryos (day 16 to 17 of gestation) and newborn mice. It is shown that fine structure of the centrosome in embryonic enterocytes differs from that found in cells of the adult mice. The centrosome of the crypt cells is more active: a greater number of cytoplasmic microtubules terminates there; mother centriole has two types of satellites; both centrioles are surrounded by fine fibrillar material. In 20% of crypt cells, replication of centrioles is observed. In the upper part of the villus of embryonic intestine, there still are two centrioles per cell, but they are located 3-7 um apart and devoid of any cytoplasmic microtubules attached to or directed toward them. In newborn mice, centrosomes of cells located at the bottom of the crypt are less active, as compared with centrosomes of embryonic cells. There are 1-3 satellites on mother centriole and few cytoplasmic microtubules closing to the centrosome. In 20% of cells from the bottom of the crypt centrioles are replicating. In the lateral part of the crypt, centrosome is inactive: centrioles are located 1-3 um apart and do not replicate. In the villus, centrioles undergo changes similar to those observed in the villus of adult mice. Centrioles loose portions of microtubule triplets. Near the top of the villus, only one centriole was found in two of 25 studied cells. In the enterocytes of the murine small intestine, centrioles are always located far from nuclei and close to the apical cell surface (1-3 um from the brush border). Centrioles never form a primary cilium and are not attached to the plasma membrane.     

Mitosis in the human embryo: the vital role of the sperm centrosome (centriole)

The pattern of sperm centrosomal (centriolar) inheritance, centrosomal replication and perpetuation during mitosis of the human embryo is reviewed with a series of electron micrographs. Embryonic cleavage involves repeated mitoses, a convenient sequence to study centriolar behaviour during cell division. After the paternal inheritance of centrioles in the human was reported (Sathananthan et al., 1991a), there has been an upsurge of centrosomal research in mammals, which largely follow the human pattern. The human egg has an inactive non-functional centrosome. The paternal centrosome contains a prominent centriole (proximal) associated with pericentriolar material which is transmitted to the embryo at fertilization and persists during sperm incorporation. Centriolar duplication occurs at the pronuclear stage (interphase) and the centrosome initially organizes a sperm aster when male and female pronuclei breakdown (prometaphase). The astral centrosome containing diplosomes (two typical centrioles) splits and relocates at opposite poles of a bipolar spindle to establish bipolarization, a prerequisite to normal cell division. Single or double centrioles occupy pivotal positions on spindle poles and paternal and maternal chromosomes organize on the equator of a metaphase spindle, at syngamy. Bipolarization occurs in all monospermic and in most dispermic ova. Dispermic embryos occasionally form two sperm asters initially and produce tripolar spindles (tripolarization). Anaphase and telophase follows producing two or three cells respectively, completing the first cell cycle. Descendants of the sperm centriole were found at every stage of perimplantation embryo development and were traced from fertilization through cleavage (first four cell cycles) to the morula and hatching blastocyst stage. Centrioles were associated with nuclei at interphase, when they were often replicating and occupied pivotal positions on spindle poles during mitosis. Sperm remnants were associated with centrioles and were found at most stages of cleavage. Centrioles were found in trophoblast, embryoblast and endoderm cells in hatching blastocysts. Pericentriolar, centrosomal material nucleated astral and spindle microtubules. Abnormal nuclear configurations observed in embryos reflect mitotic aberrations. The bovine embryo closely resembles the human embryo in centriolar behaviour during mitosis. It is concluded that the sperm centrosome is the functional active centrosome in humans and is likely the ancestor of centrioles within centrosomes in foetal and adult somatic cells. The role of the sperm centrosome in embryogenesis and male infertility is discussed, since it is of clinical importance in assisted reproduction.     

Regional differences in the pineal gland of the cotton rat, Sigmodon hispidus: light microscopic, electron microscopic, and immunohistochemical observations

Light microscopic, electron microscopic and immunohistochemical observations of the various portions of the pineal gland of the cotton rat (Sigmodon hispidus) were made. The volume of the proximal half occupied about 30% of the whole organ, and pinealocytes were slightly smaller in size in the proximal portion than elsewhere. The distal and intermediate portions contained few interstitial cells and numerous astrocytes, but the proximal portion lacked interstitial cells and had more abundant astrocytes than elsewhere. Astrocytes, which were immunoreactive for glial fibrillary acidic protein, mainly lined the pericapillary spaces in the distal and intermediate portions, but in the proximal portion these cells often surrounded isolated or groups of pinealocytes. In the distal and intermediate portions, abundant sympathetic fibers and less numerous non-sympathetic, peptidergic fibers were mainly localized in the pericapillary spaces; these fibers were sparsely distributed in the parenchyma close to interstitial cells or astrocytes. In the proximal portion, non-sympathetic fibers were scarce and sympathetic fibers were distributed abundantly and almost exclusively in the parenchyma. Most of the sympathetic fibers were adjacent to astrocytes and, occasionally, made specialized contact with them. Fenestrae in the capillary endothelium were numerous in the distal portion but absent in the proximal portion. Thus, marked differences in structure existed between the distal and proximal portions of the pineal gland of the cotton rat suggesting that both portions are functionally dissimilar. In addition, the present study indicates that the proximal portion of the cotton rat was well developed and showed morphological features similar to the deeply situated pineal glands of other mammals.     

Centrosome assembly in vitro: role of gamma-tubulin recruitment in Xenopus sperm aster formation

Centrioles organize microtubules in two ways: either microtubules elongate from the centriole cylinder itself, forming a flagellum or a cilium ("template elongation"), or pericentriolar material assembles and nucleates a microtubule aster ("astral nucleation"). During spermatogenesis in most species, a motile flagellum elongates from one of the sperm centrioles, whereas after fertilization a large aster of microtubules forms around the sperm centrioles in the egg cytoplasm. Using Xenopus egg extracts we have developed an in vitro system to study this change in microtubule-organizing activity. An aster of microtubules forms around the centrioles of permeabilized frog sperm in egg extracts, but not in pure tubulin. However, when the sperm heads are incubated in the egg extract in the presence of nocodazole, they are able to nucleate a microtubule aster after isolation and incubation with pure calf brain tubulin. This provides a two-step assay that distinguishes between centrosome assembly and subsequent microtubule nucleation. We have studied several centrosomal antigens during centrosome assembly. The CTR2611 antigen is present in the sperm head in the peri-centriolar region. gamma-tubulin and certain phosphorylated epitopes appear in the centrosome only after incubation in the egg extract. gamma-tubulin is recruited from the egg extract and associated with electron-dense patches dispersed in a wide area around the centrioles. Immunodepletion of gamma-tubulin and associated molecules from the egg extract before sperm head incubation prevents the change in microtubule-organizing activity of the sperm heads. This suggests that gamma-tubulin and/or associated molecules play a key role in centrosome formation and activity.     

A prospective randomized administration of 5''-deoxy-5-fluorouridine as adjuvant chemotherapy for hepatocellular carcinoma treated with transcatheter arterial chemoembolization

Surgical microscopy and electrophysiological techniques were used to standardize the nomenclature for the pudendal nerve and sacral plexus according to their somatic axonal composition in the male rat. We conclude that the pudendal nerve is the segment running from the L6-S1 trunk to the sacral plexus, carrying efferent fibers to the coccygeus, internal obturator, ventral and dorsal bulbospongiosus, ischiocavernosus, external anal sphincter, and external urethral sphincter muscles, and afferent fibers from the penis, prepuce, scrotum, and ventral-proximal tail. The sacral plexus is the complex formed by the bridge-like structure connecting the pudendal nerve with the lumbosacral trunk, and two nerve branches emerging from it, one innervating the proximal half of the scrotal skin, and the other innervating the muscles at the base of the penis known as the motor branch. These branches are only considered as a part of the sacral plexus because they integrate axons from both the lumbosacral trunk and pudendal nerve. The gross anatomy of the pudendal nerve and sacral plexus has a main organization that was observed in 70% of cases, whereas the remaining 30% occurred in two variants. This nomenclature is appropriate to describe the pudendal nerve and sacral plexus in studies that involve them being lesioned or electrophysiologically analysed. A main additional finding was that two large afferent branches innervate the scrotum, one the proximal half and the other the distal half. As mentioned above, the proximal branch belongs to the sacral plexus, whereas the distal branch belongs to the pudendal nerve because all its axons travel to the cord via this nerve. Since stimulation or even manipulation of the scrotal branches resulted in the secretion of semen containing spermatozoa, it is suggested that scrotal afferents are involved in some way in the ejaculatory process, a topic that deserves further research.     

Formation of alternating tiers in the optic chiasm of the chick embryo

BACKGROUND: When the fibers of the two optic nerves of the chick cross to the contralateral side at the prospective chiasmatic region, they segregate into clearly defined bundles. These bundles form horizontally oriented tiers which alternate between the right and the left optic nerve. METHODS: We have analyzed the development of these tiers qualitatively and quantitatively using light and electron microscopy between embryonic days (E) 4 and E19. RESULTS: The formation of the chiasm begins on E4. In the course of E4, tiers become visible for the first time. Their number increases rapidly until E7. Then the increase is slowed down and the final value (32 +/- 1) is approximated by E18/19. Growing axons allow one to distinguish three different segments: the growth cone, the distal, and the proximal segment. The latter originates in the perikaryon. Growth cones and distal segments are found predominantly in the ventralmost tiers. Their frequency decreases from ventral to dorsal. Proximal segments which indicate the presence of older axons appear first in the dorsal tiers and later also in more ventrally located tiers. CONCLUSION: Based on these criteria it is concluded that newly formed axons contribute primarily but not exclusively to the ventral tiers. There is a gradient of maturity of axons from ventral to dorsal whose slope becomes steeper with age until the last growth cones have arrived by E18. Thus, the formation of the chiasm corresponds to the spatiotemporal pattern of ganglion cell formation in the retina. The process of cell death of retinal ganglion cells is also seen in the chiasm but probably does not lead to a transitory diminution in the number of tiers.     

Centriole disassembly in vivo and its effect on centrosome structure and function in vertebrate cells

Glutamylation is the major posttranslational modification of neuronal and axonemal tubulin and is restricted predominantly to centrioles in nonneuronal cells (Bobinnec, Y., M. Moudjou, J.P. Fouquet, E. Desbruyères, B. Eddé, and M. Bornens. 1998. Cell Motil. Cytoskel. 39:223-232). To investigate a possible relationship between the exceptional stability of centriole microtubules and the compartmentalization of glutamylated isoforms, we loaded HeLa cells with the monoclonal antibody GT335, which specifically reacts with polyglutamylated tubulin. The total disappearance of the centriole pair was observed after 12 h, as judged both by immunofluorescence labeling with specific antibodies and electron microscopic observation of cells after complete thick serial sectioning. Strikingly, we also observed a scattering of the pericentriolar material (PCM) within the cytoplasm and a parallel disappearance of the centrosome as a defined organelle. However, centriole disappearance was transient, as centrioles and discrete centrosomes ultimately reappeared in the cell population. During the acentriolar period, a large proportion of monopolar half-spindles or of bipolar spindles with abnormal distribution of PCM and NuMA were observed. However, as judged by a quasinormal increase in cell number, these cells likely were not blocked in mitosis. Our results suggest that a posttranslational modification of tubulin is critical for long-term stability of centriolar microtubules. They further demonstrate that in animal cells, centrioles are instrumental in organizing centrosomal components into a structurally stable organelle.     

Control of spine formation by electrical activity in the adult rat cerebellum

Dendritic spines are a key structure in neuronal plasticity. Enhanced activity is commonly associated with an increase in spine size and density. Purkinje cell dendrites are characterized by a proximal and a distal compartment on which climbing fibers and parallel fibers, respectively, impinge. The proximal region has a very low spine density, whereas the distal region has a high density. Previous experiments showed that after climbing fiber deletion, Purkinje cells become hyperactive, and a large number of spines develop on the proximal dendrites. Here we show that the same hyperspiny transformation occurs in the proximal dendrites of adult Purkinje cells by depressing electrical activity with tetrodotoxin. Thus, spines in different dendritic compartments are created or maintained independently from the level of Purkinje cell-firing rate and when the afferent activity is blocked. This conclusion supports the view that spinogenesis is the expression of an intrinsic program and the two regions of the dendritic tree respond differently to activity block because of differences in the inputs that they receive. On tetrodotoxin treatment, climbing fibers become atrophic and may sprout thin collateral ramifications directed mainly toward the granular layer. All changes are reversible on tetrodotoxin removal. Therefore, Purkinje cells provide a model where spines in different compartments of the same neuron are differently regulated by the activity of their local afferents. In addition, electrical activity is also essential to maintain the full climbing fiber innervation.     

Influence of extracellular fluid volume expansion on magnesium, calcium and phosphate handling along the rat nephron

Renal tubular handling of P, Ca, Mg and Na was studied in the rat both before and during mild hypertonic NaCl loading (ECVE), using micropuncture and clearance techniques and electron microprobe analysis. Micropuncture was performed at the late proximal and early distal tubule sites. ECVE significantly increased the urinary output of all four elements. In the case of Mg, the increase was relatively small and depended on slight but statistically unsignificant inhibition of reabsorption all along the entire length of the nephron. For Ca, it depended on the inhibition of proximal reabsorption, partially compensated by increased reabsorption along the loop. For P, it depended on proximal inhibition, no important net phosphate movement occurring in the loop during both periods. Ca reabsorption was highly correlated to that of sodium along the proximal tubule and Henel's loop, Ca and Mg reabsorption were closely related to the load delivered at the beginning of the structure. These observations are compatible with the view that tubular reabsorption of Ca and Mg is concentration rather than Tm limited, and that reabsorption of Ca, unlike that of Mg, is linked to the movements of sodium. Following ECVE, the difference between early distal and urinary deliveries increased significantly for Ca and P, but not for Mg. For phosphate, this difference accounted for by 45% of the delivery at the early distal tubule site, at variance with microinjection data obtained in the rat under similar salt loading conditions, which indicated that 17% only of the phosphate distal delivery were reabsorbed along the terminal segments. This discrepancy is discussed in terms of nephron functional heterogeneity.     

Regeneration in Duchenne muscular dystrophy. Electromyographic evidence

Eight patients with Duchenne muscular dystrophy (DMD) and seven normal children of similar age were studied with a new electromyographic method for coherent displays of potentials. Five hundred motor unit potentials (MUPs) were analyzed in 20 proximal and distal muscles representing a wide spectrum of dystrophy. The progressive MUP disintegration by dropping out of muscle fibers was documented, as well as a high incidence of spontaneous fibrillation. A total of 386 late component (LC) potentials followed the 500 MUPs at consistent latencies. No LC occurred in normal children. The LCs result from motor axon sprouts innervating muscle fibers that are newly formed either by segmentation of existing muscle fibers (focal neucrosis and membrane repair) or by muscle regeneration. Axons in DMD can thus collaterally innervate additional muscle fibers. These processes must delay the onset and progression of clinical weakness in DMD patients.     

Ultrastructure of spermiogenesis and the spermatozoon of Anoplocephaloides dentata (Cestoda, Cyclophyllidea, Anoplocephalidae), an intestinal parasite of Arvicolidae rodents

Spermiogenesis in Anoplocephaloides dentata begins with the formation of a differentiation zone delimited by a ring of arched membranes. This conical area shows 2 parallel centrioles with associated anterior reduced striated roots but without an intercentriolar body. Only 1 of the centrioles develops an axoneme that grows into a cytoplasmic extension. Two crestlike bodies appear when the nucleus initiates its migration along the spermatid body. We describe for the first time at the end of spermiogenesis the formation of an apical cone before the strangulation of the ring of arched membranes. The mature spermatozoon of A. dentata is filiform, tapered at both ends, and lacks mitochondria. Its anterior extremity has an apical cone measuring about 1,400 x 350 nm and 2 crestlike bodies. Cortical microtubules are spiralled at an angle of about 30 degrees to the spermatozoon axis. The axoneme, of the 9+1 pattern of Trepaxonemata (Polycladida, Seriata, "Typhloplanoida," "Dalyellioida," and Neodermata lacks a periaxonemal sheath and does not reach the extremities of the spermatozoon. Numerous granules of electron-dense material are observed in the posterior regions of each spermatozoon. Analysis of ultrastructural features found during spermiogenesis in A. dentata corroborates the presence of striated roots associated with the centrioles in cyclophyllidean species. Moreover, the presence of striated roots is described for the first time in type IV spermiogenesis.     

Rise time and amplitude of visually elicited EPSPs of tectal neurons of the frog

Visually elicited activities in response to on-off of diffuse light and moving patterns were recorded from tectal neurons of the frog by means of perforated in vivo whole cell recording technique. Rise time histograms of EPSPs had a single or multiple peaks. Comparing the histogram patterns for each stimulus, retinal fiber classes and positions of synapses on the dendrites could be correlated. In most cells, R3 fibers made synapses on proximal dendrites, whereas R1/R2 fibers on distal dendrites, which was consistent with anatomical findings. Preliminary quantum analysis of EPSP amplitudes was performed by fitting the histograms with a binomial curve and parameters q and n were determined. The result showed that the number n of synapses on distal dendrites is larger than those on proximal dendrites.     

The role of distal and proximal interaction in infant social preference formation.

Created experimental social histories for 22 24-wk-old infants to determine the role of distal and proximal interaction in social preference formation. Each S interacted with 3 different adults during a 3-wk period. The distal interactor smiled and talked to the infant. The proximal interactor rocked and patted the infant. The neutral interactor sat by the infant without responding in any way. During 9 preference tests, infants were allowed to move in a specially adapted walker toward any of the 3 interactors. During the 3 wks, infants increased the time spent near the distal interactor. In the 3rd wk, infants looked more at the distal interactor than at the proximal interactor, and spent more time near both distal and neutral interactors than near the proximal interactor. The separate roles of distal and proximal interaction in the formation of social preferences are discussed in terms of the different contributions infants can make to distal and proximal aspects of interaction. (22 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Corticomotoneuronal postspike effects in shoulder, elbow, wrist, digit, and intrinsic hand muscles during a reach and prehension task

We used spike-triggered averaging of rectified electromyographic activity to determine whether corticomotoneuronal (CM) cells produce postspike effects in muscles of both proximal and distal forelimb joints in monkeys performing a reach and prehension task. Two monkeys were trained to perform a self-paced task in which they reached forward from a starting position to retrieve a food reward from a small cylindrical well. We compiled spike-triggered averages from 22 to 24 separate forelimb muscles at both proximal (shoulder, elbow) and distal (wrist, digits, intrinsic hand) joints. Of 174 cells examined, 112 produced postspike effects in at least one of the target muscles. Of those cells, 45.5% produced postspike effects in both proximal and distal forelimb muscles. A nearly equal number (44.7%) produced postspike effects in distal muscles only, whereas a clear minority (9.8%) produced postspike effects in only proximal muscles. The majority of CM cells (71.4%) produced effects in two or more muscles, with an average muscle field of 3.1 +/- 2.1 (mean +/- SD) for facilitation plus suppression. Of 345 postspike effects identified, 70.7% were facilitation effects and 29.3% were suppression effects. The large majority of effects (72.2%) were in distal muscles. When averaged by joint, the latency and peak magnitude of postspike facilitation showed a stepwise increase from proximal to distal joints. The results of this study show that the majority of CM cells engaged in coordinated forelimb reaching movements facilitate and/or suppress muscles at multiple joints, including muscles at both proximal and distal joints. The results also show that CM cells make more frequent and more potent terminations in motoneuron pools of distal compared with proximal muscles.