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Unidirectional recruitment between MeCP2 and KSHV-encoded LANA revealed by CRISPR/Cas9 recruitment assay.

Kaposi's sarcoma-associated herpesvirus (KSHV, HHV-8) is associated with several human malignancies. During latency, the viral genomes reside in the nucleus of infected cells as large non-integrated plasmids, known as episomes. To ensure episome maintenance, the latency protein LANA tethers the viral episomes to the cell chromosomes during cell division. Directional recruitment of protein complexes is critical for the proper function of many nuclear processes. To test for recruitment directionality between LANA and cellular proteins, we directed LANA via catalytically inactive Cas9 (dCas9) to a repeat sequence to obtain easily detectable dots. Then, the recruitment of nuclear proteins to these dots can be evaluated. We termed this assay CRISPR-PITA for Protein Interaction and Telomere Recruitment Assay. Using this protein recruitment assay, we found that LANA recruits its known interactors ORC2 and SIN3A. Interestingly, LANA was unable to recruit MeCP2, but MeCP2 recruited LANA. Both LANA and histone deacetylase 1 (HDAC1) interact with the transcriptional-repression domain (TRD) and the methyl-CpG-binding domain (MBD) of MeCP2. Similar to LANA, HDAC1 was unable to recruit MeCP2. While heterochromatin protein 1 (HP1), which interacts with the N-terminal of MeCP2, can recruit MeCP2. We propose that available interacting domains force this recruitment directionality. We hypothesized that the tandem repeats in the SunTag may force MeCP2 dimerization and mimic the form of DNA-bound MeCP2. Indeed, providing only the tandem epitopes of SunTag allows LANA to recruit MeCP2 in infected cells. Therefore, CRISPR-PITA revealed the rules of unidirectional recruitment and allowed us to break this directionality.

Humans

Respiratory effects of recruitment maneuvers according to lung recruitability assessed by electrical impedance tomography in patients with acute respiratory distress syndrome.

Recruitment maneuvers (RM) can improve oxygenation in patients with acute respiratory distress syndrome (ARDS), but their physiological effects depend on lung recruitability. This secondary analysis of a randomized controlled trial (RCT) evaluated oxygenation, respiratory mechanics, regional ventilation, and cardiorespiratory adverse events responses to a RM followed by electrical impedance tomography (EIT)-guided PEEP titration, using EIT to assess lung recruitability. In this study, fifty patients with moderate-to-severe ARDS underwent a stepwise RM followed by individualized PEEP titration guided by EIT. Lung recruitability was determined using the collapse index at PEEP 6 cmH&#x2082;O (CLPEEP6), defined as the proportion of collapsed lung at this PEEP level. Patients were classified into high- and low-recruitability groups based on median CLPEEP6 values. Oxygenation (PaO&#x2082;/FiO&#x2082;), static compliance (Cstat), driving pressure (Pdriv), regional ventilation distribution, and cardiorespiratory adverse events were compared before and after RM, during subsequent individualized EIT-guided PEEP titration. In patients with high recruitability (CLPEEP6&#x2009;>&#x2009;12.5), the PaO&#x2082;/FiO&#x2082; ratio and Cstat increased significantly after RM (PaO&#x2082;/FiO&#x2082;: 100.8&#x2009;&#xb1;&#x2009;30.3 vs. 125.4&#x2009;&#xb1;&#x2009;38.3&#xa0;mmHg, p&#x2009;<&#x2009;0.05; Cstat: 21.5&#x2009;&#xb1;&#x2009;5.8 vs. 28.0&#x2009;&#xb1;&#x2009;7.0&#xa0;mL/cmH&#x2082;O, p&#x2009;<&#x2009;0.001), Pdriv decreased (19.1&#x2009;&#xb1;&#x2009;3.5 vs. 15.6&#x2009;&#xb1;&#x2009;3.2 cmH&#x2082;O, p&#x2009;<&#x2009;0.001). EIT demonstrated a posterior redistribution of ventilation after RM. In contrast, patients with low recruitability (CLPEEP6&#x2009;&#x2264;&#x2009;12.5) showed no significant mechanical or oxygenation improvement and transient hypotension, arrhythmia, and desaturation appeared numerically more common in this group. No barotrauma or cardiac arrest occurred, and ICU mortality was similar between groups. A strategy combining a RM with subsequent individualized EIT-guided PEEP titration was associated with improved oxygenation and lung mechanics in patients with high lung recruitability, whereas patients with low recruitability showed limited physiological benefit, with cardiorespiratory adverse events appearing numerically more frequent. The EIT-derived CLPEEP6 index represents a feasible and clinically applicable https://clinicaltrials.gov/study/NCT06733168.

Humans

Recruitment of effector lymphocytes by initiator lymphocytes. Recruited lymphocytes are immunospecific and include graft-versus-host-reactive lymphocytes.

We have shown previously that initiator T lymphocytes (ITL), sensitized in vitro against fibroblast antigens, recruit effector T cells in vivo. After injection into hind footpads of syngeneic recipients, sensitized ITL migrated to the draining popliteal lymph nodes (PLN) and activated a trapping mechanism by which circulating lymphocytes were recruited in the PLN. This paper reports experiments designed to test the immunospecificity of these recruited T lymphocytes (RTL). We found that immunospecific RTL were depleted from other lymphoid organs during recruitment in the PLN. However, immunospecific ITL were not depleted from spleens during PLN recruitment. Thus ITL and RTL are functionally distinguishable. We show that specific GVH reactive lymphocytes were also lost from spleens and distal lymph nodes during trapping of RTL in the PLN. Thus, the trapping phase of the recruitment response is immunospecific, as are the sensitization and effector phases. The trapped RTL are antigen-specific, and include the pool of GVH-reactive-lymphocytes committed to the same alloantigen. Thus, it appears that GVH-reactive cells respond to syngeneic ITL sensitized against allogeneic fibroblasts.

Animals

Patient recruitment: other examples of recruitment problems and solutions.

Plans to recruit patients for a clinical trial must begin early and must utilize a variety of workable techniques if patients are to be acquired at an acceptable rate. Publicity campaigns must be directed toward winning the confidence and good will of the public and of interested physicians.

Clinical Trials as Topic

Strategies to improve recruitment to randomised trials.

BACKGROUND: Recruiting participants to randomised controlled trials (RCTs) is challenging. Identifying effective recruitment strategies would benefit health research: poor recruitment leads to underpowered trials, reducing the reliability of findings and increasing the risk of wasted resources, ethical concerns, and trial failure. Evidence to inform recruitment strategies is increasingly generated through Studies Within A Trial (SWATs), which are methodological studies embedded within host RCTs. This is an update of a review last published in 2018. OBJECTIVES: Primary: to quantify the effects of strategies to improve recruitment of participants to RCTs. Secondary: to evaluate recruitment strategies' cost-effectiveness and impact on retention, and the equity, diversity, and inclusion (EDI) characteristics of recruited participants. SEARCH METHODS: We used MEDLINE, Embase, and six other databases to identify the studies included in the review. We also sought unpublished recruitment SWATs through social media and targeted email dissemination to trial methodology networks. The latest search date was 16 February 2023. SELECTION CRITERIA: We included randomised SWATs evaluating trial recruitment strategies embedded in healthcare and non-healthcare trials. We excluded quasi-randomised, hypothetical, questionnaire-only, retention-only, or clinician incentive studies. DATA COLLECTION AND ANALYSIS: Primary outcome: proportion of eligible participants or centres recruited. SECONDARY OUTCOMES: cost-effectiveness, retention rates, and EDI characteristics of included participants. We conducted random-effects meta-analysis for strategies evaluated in at least two studies; otherwise, we synthesised results narratively. We reported effects as risk differences (RDs) with 95% confidence intervals (CIs), and assessed between-trial heterogeneity. We used GRADE to assess the certainty of evidence for the primary outcome. We expressed cost-effectiveness as the incremental cost per additional participant recruited in pounds sterling (GBP). MAIN RESULTS: We identified 91 eligible studies (53 new to this update), providing 94 comparisons and involving at least 176,747 participants. Eighty-one studies involved strategies aimed at trial participants, while 10 evaluated strategies aimed at recruiters. All were healthcare studies. We found 65 recruitment strategies; 49 were evaluated in a single study. Only five strategies were supported by high-certainty evidence according to GRADE criteria, and we focus on these strategies in the summary below. Open-label trials versus blinded, placebo trials. Open-label trials recruited more participants than blinded trials (RD 10%, 95% CI 8% to 12%; 3 studies, 9004 participants), corresponding to approximately 10 additional participants per 100 approached. The studies involved mostly women in the UK and Estonia. No cost or retention data were reported. Telephone reminder versus no telephone reminder. Telephone reminders to people who did not respond to an initial postal invitation boosted recruitment by 6% (95% CI 3% to 9%; 2 studies, 1450 participants), in trials with low underlying recruitment (we are less certain for trials with over 10% recruitment). The studies involved people with a mean age of 58 years in Canada and Norway. No cost or retention data were reported. Recruitment primer letter versus no letter. Pre-recruitment letters and leaflets designed to encourage participation made little or no difference to recruitment (absolute improvement 1%, 95% CI -1% to 2%; 2 studies, 5376 participants), and were associated with increased costs compared to not sending a primer (incremental cost: GBP 2.08). The studies involved mostly older white people in the UK and Ireland. Multimedia information via a digital link/QR code plus paper participant information leaflet (PIL) versus paper PIL alone. This made little or no difference to recruitment (absolute improvement 0%, 95% CI -1% to 1%; 7 studies, 11,612 participants) and retention (absolute improvement 0%, 95% CI -2% to 3%; 5 studies, 7403 participants), and increased costs compared to not including multimedia information (incremental cost: GBP 0.78). The studies involved people in the UK. Optimised, user-tested PIL versus standard PIL. Optimising participant information leaflets (e.g. through user-testing the leaflet with the target population to shape its content, format, and appearance) made little or no difference to recruitment: absolute improvement was 0% (95% CI 0% to 1%; 6 studies, 27,805 participants). The studies involved people in the UK. Only one study reported EDI data; participants were mostly older women. No cost or retention data were reported. We had moderate-certainty evidence for 13 other strategies; confidence was often reduced because the results came from single studies. Seven strategies involved changes to how potential participants received information; four involved changes to trial conduct; one targeted the recruiter or recruitment site; and one tested non-monetary incentives. We had much less confidence in the other 47 comparisons because the studies had design flaws, were single studies, or had very uncertain results. Costs were reported in only 17 of 91 studies. Strategy impact on retention was reported in 15 studies. All but one study (99%) were from high-income countries. The most reported demographics were age (49 studies), sex (32 studies), gender (27 studies), and education level (16 studies). AUTHORS' CONCLUSIONS: The evidence on strategies to improve trial recruitment remains broad but lacks depth. Of 65 strategies evaluated, only five were supported by high-certainty evidence. Open-label trial designs and telephone reminders to non-responders increased recruitment, while optimised participant information leaflets, recruitment primer letters, and multimedia information provided alongside a paper participant information leaflet had little or no effect. Reporting of participant characteristics was poor, limiting assessment of equity, diversity, and inclusion across most studies. Evidence is heavily skewed toward high-income countries. Future research must prioritise evaluations in low-to-middle-income settings and consistently report cost, retention, and EDI outcomes. We strongly urge the methodology research community to strengthen the evidence base by prioritising replications of existing strategies over the development and testing of new ones. FUNDING: National Institute for Health and Care Research (Advanced Fellowship, Adwoa Parker, reference:NIHR302256). Health Research Board, Republic of Ireland, Evidence Synthesis Ireland (grant ESI-2021-001) REGISTRATION: This review updates an earlier Cochrane review, which was first published in 2002 and subsequently updated in 2007, 2010, and 2018. Previous versions of the review and their protocols are available at: https://doi.org/10.1002/14651858.MR000013.pub2 https://doi.org/10.1002/14651858.MR000013.pub3 https://doi.org/10.1002/14651858.MR000013.pub4 https://doi.org/10.1002/14651858.MR000013.pub5 https://doi.org/10.1002/14651858.MR000013.pub6.

Randomized Controlled Trials as Topic

Recruitment and discharge frequency of phrenic motoneurones during inspiration.

The discharges of 107 phrenic motor axons were recorded from cats under chloralose-urethane anaesthesia with spinal cords transected at T1 or with intact neuraxis. During inspiratory occlusions in spinal cats, each motoneurone was recruited at a mouth pressure constant at a given end tidal CO2; no motoneurone was recruited at a pressure greater than 70% of maximum. In eupnoea (32.3 torr CO2) 73% of motoneurones were recruited during the first 30% of inspiration; during CO2 rebreathing (60.8 torr CO2), 89% were recruited in the first 30% of inspiration. Neurones recruited earlier in inspiration had a lower onset frequency than later recruited units; all increased instantaneous frequency in a linear relation to pressure. Early recruited units showed a smaller increase in frequency per unit change in pressure than did later recruited units. During CO2 rebreathing, mean and peak frequencies increased on average 0.92 and 1.78 spikes.sec(-1) (%CO2)(-1), respectively, these increases being significantly less for early than for late recruited neurones. The data show that a stable order of recruitment of phrenic motoneurones exists during inspiration, the excitability of each motoneurone likely determining its time of recruitment. Above threshold, later recruited motoneurones are more 'sensitive' to a change in input. Recruitment of motoneurones is responsible for pressure generation at the start of inspiration and increase in discharge frequency (rate coding) is the dominant mechanism in the second half of inspiration.

Airway Obstruction

Weathering the storm: Most maternal and environmental drivers of individual reproductive success do not scale up to population recruitment in a large herbivore.

Population growth depends upon individual survival and reproduction, but do drivers of individual reproductive success scale up to population recruitment? Factors affecting individuals may have little effect on population dynamics if individuals within a population experience different conditions. When seasonal resource availability is unpredictable and breeding season long, average conditions over a breeding cycle may poorly reflect the environment experienced by many individuals. We compared the drivers of individual reproductive success and population recruitment in an asynchronously breeding large herbivore, the eastern grey kangaroo (Macropus giganteus). We analysed 18&#x2009;years of individual-based data using multivariate hierarchical Bayesian models to first identify the causal mechanisms relating population density, environmental conditions and maternal traits to individual success. We then assessed whether the drivers of individual reproductive success scaled up to determine population recruitment. Most maternal and environmental covariates strongly influenced individual reproductive success, with distinct effects on juvenile survival before and after pouch exit. Maternal traits had a greater influence in the pouch, whereas environmental conditions became increasingly important once young exited the pouch. Most drivers of individual reproductive success did not affect population recruitment. Recruitment increased with population density and mean body condition of adult females. Weather harshness had a weak positive effect on recruitment, which appeared independent of female age structure, previous recruitment or forage. Most drivers of individual reproductive success did not scale up to population recruitment. Birth asynchrony could buffer population recruitment against environmental variation such that variables affecting individual reproduction have little impact at the population level. Large herbivores that reproduce asynchronously may therefore be more resilient to environmental variability than synchronous breeders.

Bayesian modelling

The relationship between the rate of rise of isometric tension and motor unit recruitment in a human forearm muscle.

The recruitment properties of single motor units from the human extensor indicis muscle were investigated during voluntary isometric contractions of different rate of rise but equal amplitude. Both the electrical and the contractile events associated with recruitment were analyzed. The threshold force of recruitment (measured as the total muscle force at firing onset) decreased with increasing rate of rise of isometric tension. This was consistently found for all units. Differences between low and high threshold units indicating a preferential tonic or phasic mode of activation were not observed. The contractile events associated with recruitment were analyzed analoguously to the electrical events. For this purpose, muscle force was measured at the time of the first twitch as it was measured at the time of the first spike. This separate measurement of the electrical and mechanical recruitment of a unit is necessary, because during a change of muscle force, force is different at firing onset and during the subsequent twitch contraction. Muscle tension at the time of the peak of the first twitch contraction was calculated from measurements of the twitch concentration time of the single motor units. In contrast to firing onset, the peak of the first twitch of a motor unit occurs at approximately the same muscle tension no matter how fast the contraction is performed. This is the consequence of the result that the average decrease of the threshold force of recruitment at successively faster contraction has the same value as the corresponding increase of total muscle force during the mean contraction time of the motor units. On the basis of this precise matching between these two changes, the mechanical recruitment of motor units occurs at approximately the same force level irrespective of the rate of rise of tension.

Electric Stimulation

Recruitment for the Coronary Primary Prevention Trial.

A preliminary account of some aspects of recruitment for the Coronary Primary Prevention Trial is provided. Extension of recruitment sources to include various forms of community screening resulted in more rapid acquisition of potential participants than was possible when recruitment was restricted to the more traditional sources of physician and laboratory referral. Extended recruitment was supported by augmented data feedback and by the development of the job of recruitment coordinator at each participating clinic. The identification of delays in reaching the maximal recruitment rate from particular sources points to the need for detailed planning of a recruitment campaign before large-scale clinical trials are begun.

Clinical Trials as Topic

Thalamo-cortical systems regulating spindle bursts and recruiting responses. II. Effect of thalamic lesions.

The effects of thalamic lesions on recruiting responses and spindle bursts have been studied in 12 cats immobilized with gallamine. Recruiting responses were elicited by stimulation of the midline thalamic nuclear complex. Spindle bursts were produced by lesions in the mesencephalic tegmentum. Recruiting responses and spindle bursts were recorded from the cortex and thalamus. The effect of thalamic lesions on such spontaneous and induced potentials was evaluated by making lesions anteriorly in the rostral pole of the thalamus, medially in the midline thalamic nuclear group, and laterally in the primary and association nuclei of the thalamus. 1. Small, anterior, thalamic lesions blocked spindle bursts and recruiting responses both at the cortex and in the thalamus. 2. Massive, medial, thalamic lesions blocked spindle bursts in the suprasylvian cortex, but did not affect either frontal cortex spindle bursts or frontal and thalamic recruiting responses induced by stimulation of N. ventralis anterior. 3. Massive, lateral, thalamic lesions including specific relay and association nuclei, failed to block spontaneous spindles and recruiting responses in frontal cortex and midline thalamic nuclei. It has been postulated that the rostral pole of the thalamus constitutes an important part of the thalamic portion of a thalamo-orbitocortical system which mediates and regulates electrocortical synchronization as manifested in spindle bursts, recruiting responses and synchronized wave-like potentials.

Animals

Capillary recruitment during airway hypoxia: role of pulmonary artery pressure.

Hypoxia has been shown to cause an increased number of pulmonary capillaries to be perfused. Changes in cardiac output and left atrial pressure have been previously ruled out as causes of this capillary recruitment. Increased pulmonary vein pressure and increased pulmonary artery pressure remain as two potential mechanisms. To differentiate between these two possible causes, we measured pulmonary artery and vein pressures with directly placed catheters and capillary recruitment with in vivo microscopy. During isocapnic hypoxia pulmonary artery pressure doubled, observed capillary recruitment increased fivefold, and pulmonary vein pressure remained constant. When the vasodilator prostaglandin E1 was infused during hypoxia, pulmonary artery pressure and capillary recruitment fell to control values and pulmonary vein pressure remained constant. Since capillary recruitment correlated with pulmonary artery pressure in each dog, but not with pulmonary vein pressure, we conclude that arterial, not venous, constriction is the probable cause of this recruitment.

Animals

Recruitment and firing rate modulation of motor unit tension in a small muscle of the cat's foot.

Maintained contractions were elicited in the first deep lumbrical muscle of the cat's foot by electrical stimulation of the contralateral motor cortex or, reflexly, by pinching of the foot pad. The discharges of all significant motor units of the muscle were monitored by electromyography, and contractions of the various motor units were observed in isometric recordings of muscle tension. Over a wide range, muscle tension could be enhanced by an increased intensity of pad pinching or cortical stimulation. This increase in muscle tension was caused by a recruitment of new motor units as well as by an increase in the firing rate of already active motor units. The latter mechanism was clearly of great importance. Pad pinching or cortical stimulation could sometimes cause the muscle to produce a tension close to that of a maximum tetanic contraction. This was several times greater than the mean tension that would have been caused by motor unit recruitment alone (i.e. by the motor units firing at their minimum steady rate). Cortical stimulation as well as pad pinching commonly recruited weak units more easily than stronger ones of the same muscle. The recruitment order obtained in response to pad pinching often differed, however, in various details from the recruitment caused by cortical stimulation.

Animals

[SISI or differential limen test - tests for recruitment (author's transl)].

The detection of recruitment is based on investigating change of loudness with change of intensity. The evaluation and further differential diagnosis of inner ear damage is, for instance, a necessity in an expert opinion of an ENT specialist. Which of the numerous methods of detecting recruitment can be recommended for the specialists practice. Over 215 patients with inner ear damage were tested for recruitment with different test methods. Results showed that SISI tests were performed with more accuracy than differential limen tests. Differences of various recruitment methods of our collective are investigated and advantages and disadvantages discussed. The recommendation for the specialists practice would be to perform 1. the "limits of intensity" (threshold of hearing and loudness discomfort), 2. the supraliminal noise-audiometry Langenbeck and 3. the SISI test. They seem to be most effectful, exact, reproducible and easy to perform test-methods for detecting recruitment.

Audiometry

[Investigations on the vestibular recruitment (author's transl)].

Searching for some kind of vestibular recruitment that might be similar to the audiological recruitment as defined by Fowler, a caloric test was done at 44 degrees, 33 degrees, 30 degrees, 26 degrees, 17 degrees. The parameter of vestibular response was the maximum slow component speed, In total 36 Menière-patients were electronystagmographically examined. Out of the 36 only 16 were analyzed further for the question of vestibular recruitmen, who showed strictly unilateral lesions, who were attack-free since some months and who showed clear audiological recruitment at four different recruitment tests. The following results were obtained: 1. The values obtained for the sound side of Menière-patients correspond to values described as normal in literature. 2. The affected side showed less reaction at all temperatures, in no case there was a tendency to recruitment. 3. With considerable loss of excitability, a "flat curve" was resulting, indicating decruitment.

Audiometry

Leukocyte recruitment to airways by aldehyde-carbon combinations that mimic cigarette smoke.

Exposure of Syrian golden hamsters to formaldehyde (3 to 250 p.p.m.) evaporated onto carbon (21 to 805 mg. per cu. m.) recruited polymorphonuclear (PMN) leukocytes to the epithelium of tracheas and intrapulmonary airways which peaked at 24 to 48 hours. Acrolein (less than 6 p.p.m.) on carbon (593 mg. per cu. m.) caused PMN recruitment which was maximal at 12 hours. The vapor phase of cigarette smoke produced PMN leukocyte recruitment of the same magnitude and with the same time course. In contrast, exposure to formaldehyde at doses of 2 to 250 p.p.m. and acrolein at 6 p.p.m. was cytotoxic to airway cells and caused prompt and delayed exfoliation but no recruitment of PMN leukocytes. There was no difference in cytotoxicity when carbon was present. Leukocyte recruitment occurred only when carbon was present, either given simultaneously wiht aldehydes or with adsorbed aldehydes. Thus, aldehyde vapor simulates the cytotoxic effects of particle-free cigarette smoke vapor. Of greater significance is the finding that an aldehyde, formaldehyde or acrolein, inhaled adsorbed on carbon or simultaneously with carbon to hamster airways is chemotactic for PMN leukocytes just as is the vapor phase of cigarette smoke when given simultaneously with carbon particles.

Acrolein

Thalamo-cortical projections for recruiting responses and spindling-like responses in the parietal cortex.

1. The thalamic neurones sending their axons to the parietal association cortex (middle suprasylvian gyrus) and receiving monosynaptic excitation from the cerebellar (interpositus or lateral) nucleus were recorded with microelectrodes extracellularly and intracellularly around the anterior ventral (VA) nucleus of the thalamus in cats. Such thalamic neurones are known to carry exclusively the impulses responsible for superficial thalamo-cortical (T-C) responses in the parietal cortex, being called superficial T-C neurones (see Sasaki et al., 1972a, b). 2. Repetitive (6--9/sec) stimulation of the centrum medianum-parafascicular complex (CM) or the intralaminar nuclei (IL) of the thalamus elicited grouped spike discharges of the neurone in synchronization with the recruiting responses in the parietal cortex. The grouped discharges usually preceded the respective cortical responses by several milliseconds. Numbers of the spikes in the grouped discharges increased and decreased as the recruiting responses waxed and waned on the repetitive stimulation. 3. The superficial T-C neurones also showed similar grouped discharges in synchronization with spindling-like, surface-negative cortical responses which occurred spontaneously or were evoked by single thalamic stimulation. 4. It was concluded that the superficial T-C neurons can convey impulses for recruiting responses and spindling-like responses from the thalamus directly to the cerebral cortex. They are supposed to constitute the final T-C pathway of the neuronal circuits of the recruiting system, i.e., non-specific T-C projection system.

Animals

Disturbances in voluntary recruitment order of low and high frequency motor units on blockades of proprioceptive afferent activity.

The voluntary recruitment order of anterior tibial motor units was studied in electromyographic recordings using both low and high impedance electrodes. It was shown in previous papers, that the recruitment order in sustained voluntary contraction is normally stable and that motor units with low thresholds discharge at low, regular frequencies and that motor units with high thresholds discharge at higher and more irregular frequencies. In this paper the consequences for the discharge properties of reduced afferent inflow in sustained voluntary contraction were studied, using partial ischemic or lidocain blockades of the muscle nerve and local cooling of the muscle belly. 1. The recruitment order of certain low and certain high frequency units was reversed. 2. The muscle tension and sense of effort necessary for tonic firing were increased for certain low frequency units but decreased for certain high frequency units. 3. On severe reduction of the afferent inflow, the differences in discharge pattern between low and high frequency units were decreased. It is concluded that the recruitment of low frequency units before high frequency units in sustained voluntary contraction is partly due to proprioceptive afferent activity favouring low frequency units and disfavouring high frequency units. It is also concluded that the afferent inflow is involved in restricting certain units to low frequency discharge and others to high frequency discharge.

Action Potentials