Key aspects of phrenic motoneuron and diaphragm muscle development during the perinatal period.

At the time of birth, respiratory muscles must be activated to sustain ventilation. The perinatal development of respiratory motor units (comprising an individual motoneuron and the muscle fibers it innervates) shows remarkable features that enable mammals to transition from in utero conditions to the air environment in which the remainder of their life will occur. In addition, significant postnatal maturation is necessary to provide for the range of motor behaviors necessary during breathing, swallowing, and speech. As the main inspiratory muscle, the diaphragm muscle (and the phrenic motoneurons that innervate it) plays a key role in accomplishing these behaviors. Considerable diversity exists across diaphragm motor units, but the determinant factors for this diversity are unknown. In recent years, the mechanisms underlying the development of respiratory motor units have received great attention, and this knowledge may provide the opportunity to design appropriate interventions for the treatment of respiratory disease not only in the perinatal period but likely also in the adult.     

Phrenic motoneuron morphology during rapid diaphragm muscle growth.

In the adult rat, there is a general correspondence between the sizes of motoneurons, motor units, and muscle fibers that has particular functional importance in motor control. During early postnatal development, after the establishment of singular innervation, there is rapid growth of diaphragm muscle (Dia(m)) fibers. In the present study, the association between Dia(m) fiber growth and changes in phrenic motoneuron size (both somal and dendritic) was evaluated from postnatal day 21 (D21) to adulthood. Phrenic motoneurons were retrogradely labeled with fluorescent tetramethylrhodamine dextran (3,000 MW), and motoneuron somal volumes and surface areas were measured using three-dimensional confocal microscopy. In separate animals, phrenic motoneurons retrogradely labeled with choleratoxin B-fragment were visualized using immunocytochemistry, and dendritic arborization was analyzed by camera lucida. Between D21 and adulthood, Dia(m) fiber cross-sectional area increased by approximately 164% overall, with the growth of type II fibers being disproportionate to that of type I fibers. There was also substantial growth of phrenic motoneurons ( approximately 360% increase in total surface area), during this same period, that was primarily attributable to an expansion of dendritic surface area. Comparison of the distribution of phrenic motoneuron surface areas between D21 and adults suggests the establishment of a bimodal distribution that may have functional significance for motor unit recruitment in the adult rat.     

Phrenic and external intercostal motoneuron activity during progressive asphyxia

Phrenic and external intercostal motoneuron activities were compared during progressive asphyxia induced by the interruption of artificial ventilation in the pentobarbital-urethan-anesthetized, gallamine-paralysed rabbit. The relative augmentation of inspiratory activity of the T1-T4 external intercostal nerves was significantly greater than that of the phrenic nerve during asphyxic hyperpnea. This was associated with a greater recruitment of intercostal than of phrenic motoneurons, particularly late in the hyperpneic phase immediately before the period of asphyxic apnea. However, peak and average discharge frequencies developed by intercostal motoneurons (n = 20) were only approximately 60% of those of the phrenic motoneurons (n = 28). Gasping respiration terminated the apneic period and was associated with a further intense recruitment of intercostal though not of phrenic motoneurons, but discharge frequencies developed by the intercostal motoneurons remained approximately 60% of those of the phrenic motoneurons. The instantaneous frequency profiles generated by the motoneurons often exhibited progressive changes during the terminal stages of hyperpnea (reduction in inspiratory duration and duty cycle and increases in inspiratory slope and discharge frequencies) such that much of the character of gasping respiration became evident before the apnea. Such smooth transitional sequences do not obviate the existence of an "independent gasping center" but do require that such a proposed center at least possess the capacity for interaction with those sites responsible for the generation of eupneic and hyperpneic respiration.     

Phrenic motoneuron discharge during sustained inspiratory resistive loading

Iscoe, Steve. Phrenic motoneuron discharge duringsustained inspiratory resistive loading. J. Appl.Physiol. 81(5): 2260-2266, 1996.I determinedwhether prolonged inspiratory resistive loading (IRL) affects phrenicmotoneuron discharge, independent of changes in chemical drive. Inseven decerebrate spontaneously breathing cats, the discharge patternsof eight phrenic motoneurons from filaments of one phrenic nerve weremonitored, along with the global activity of the contralateral phrenicnerve, transdiaphragmatic pressure, and fractional end-tidalCO₂ levels. Discharge patterns during hyperoxic CO₂ rebreathingand breathing against an IRL (2,500-4,000cmH₂O ^· l^{1 ·} s)were compared. During IRL, transdiaphragmatic pressure increased andthen either plateaued or decreased. At the highest fractional end-tidalCO₂ common to both runs,instantaneous discharge frequencies in six motoneurons were greaterduring sustained IRL than during rebreathing, when compared at the sametime after the onset of inspiration. These increased dischargefrequencies suggest the presence of a load-induced nonchemical drive tophrenic motoneurons from unidentified source(s).

     

Contractile and fatigue properties of the rat diaphragm musculature during the perinatal period.

The following two hypotheses regarding diaphragm contractile properties in the perinatal rat were tested. First, there is a major transformation of contractile and fatigue properties during the period between the inception of inspiratory drive transmission in utero and birth. Second, the diaphragm muscle properties develop to functionally match changes occurring in phrenic motoneuron electrophysiological properties. Muscle force recordings and intracellular recordings of end-plate potentials were measured by using phrenic nerve-diaphragm muscle in vitro preparations isolated from rats on embryonic day 18 and postnatal days 0-1. The following age-dependent changes occurred: 1) twitch contraction and half relaxation times decreased approximately two- and threefold, respectively; 2) the tetanic force levels increased approximately fivefold; 3) the ratio of peak twitch force to maximum tetanic force decreased 2.3-fold; 4) the range of forces generated by the diaphragm in response to graded nerve stimulation increased approximately twofold; 5) the force-frequency curve was shifted to the right; and 6) the propensity for neuromuscular transmission failure decreased. In conclusion, the diaphragm contractile and phrenic motoneuron repetitive firing properties develop in concert so that the full range of potential diaphragm force recruitment can be utilized and problems associated with diaphragm fatigue are minimized.     

Voltage-sensitive calcium currents and their role in regulating phrenic motoneuron electrical excitability during the perinatal period

This study examined the ontogeny of voltage-sensitive calcium conductances in rat phrenic motoneurons (PMNs) and their role in regulating electrical excitability during the perinatal period. Specifically, we studied the period spanning from embryonic day (E)16 through postnatal day (P)1, when PMNs undergo fundamental transformation in their morphology, passive properties, ionic channel composition, synaptic inputs, and electrical excitability. Low voltage-activated (LVA) and high voltage-activated (HVA) conductances were measured using whole cell patch recordings utilizing a cervical slice-phrenic nerve preparation from perinatal rats. Changes between E16 and P0-1 included the following: an approximately 2-fold increase in the density of total calcium conductances, an approximately 2-fold decrease in the density of LVA calcium conductances, and an approximately 3-fold increase in the density of HVA conductances. The elevated expression of T-type calcium channels during the embryonic period lengthened the action potential and enhanced electrical excitability as evidenced by a hyperpolarization-evoked rebound depolarization. The reduction of LVA current density coupled to the presence of a hyperpolarizing outward A-type potassium current had a critical effect in diminishing the rebound depolarization in neonatal PMNs. The increase in HVA current density was concomitant with the emergence of a calcium-dependent "hump-like" afterdepolarization (ADP) and burst-like firing. Neonatal PMNs develop a prominent medium-duration afterhyperpolarization (mAHP) as the result of coupling between N-type calcium channels and small conductance, calcium-activated potassium channels. These data demonstrate that changes in calcium channel expression contribute to the maturation of PMN electrophysiological properties during the time from the commencement of fetal inspiratory drive to the onset of continuous breathing at birth.     

Voltage‐sensitive calcium currents and their role in regulating phrenic motoneuron electrical excitability during the perinatal period

This study examined the ontogeny of voltage‐sensitive calcium conductances in rat phrenic motoneurons (PMNs) and their role in regulating electrical excitability during the perinatal period. Specifically, we studied the period spanning from embryonic day (E)16 through postnatal day (P)1, when PMNs undergo fundamental transformation in their morphology, passive properties, ionic channel composition, synaptic inputs, and electrical excitability. Low voltage‐activated (LVA) and high voltage‐activated (HVA) conductances were measured using whole cell patch recordings utilizing a cervical slice‐phrenic nerve preparation from perinatal rats. Changes between E16 and P0–1 included the following: an ≈2‐fold increase in the density of total calcium conductances, an ≈2‐fold decrease in the density of LVA calcium conductances, and an ≈3‐fold increase in the density of HVA conductances. The elevated expression of T‐type calcium channels during the embryonic period lengthened the action potential and enhanced electrical excitability as evidenced by a hyperpolarization‐evoked rebound depolarization. The reduction of LVA current density coupled to the presence of a hyperpolarizing outward A‐type potassium current had a critical effect in diminishing the rebound depolarization in neonatal PMNs. The increase in HVA current density was concomitant with the emergence of a calcium‐dependent “hump‐like” afterdepolarization (ADP) and burst‐like firing. Neonatal PMNs develop a prominent medium‐duration afterhyperpolarization (mAHP) as the result of coupling between N‐type calcium channels and small conductance, calcium‐activated potassium channels. These data demonstrate that changes in calcium channel expression contribute to the maturation of PMN electrophysiological properties during the time from the commencement of fetal inspiratory drive to the onset of continuous breathing at birth. © 2001 John Wiley & Sons, Inc. J Neurobiol 46: 231–248, 2001     

Phrenic motoneuron firing rates before, during, and after prolonged inspiratory resistive loading

Road, J. D., and A. M. Cairns. Phrenic motoneuronfiring rates before, during, and after prolonged inspiratory resistive loading. J. Appl. Physiol. 83(3):776-783, 1997.Phrenic motoneuron firing rates during briefinspiratory resistive loading (IRL) are high, and nearly all themotoneurons are recruited. Diaphragmatic fatigue has been difficult todemonstrate during IRL. Furthermore, evidence from studies in limbmuscles has shown variable motoneuron responses to prolongedhigh-intensity loads. We studied phrenic motoneuron firing ratesbefore, during, and after prolonged IRL in anesthetized rabbits. Of 117 phrenic axons, only 2 axons were not recruited; 41 axons were silentduring unloaded breathing but were recruited at higher loads. Silentaxons showed a more rapid increase in firing rate as the loadincreased. Phrenic motoneuron firing rates increased throughout theperiod of loading, whereas airway pressure swings did not. Afterprolonged IRL, higher motoneuron firing rates were needed during briefloads to produce the same airway pressure. No evidence of a decline inmotoneuron firing rates was seen at any point. We conclude that therespiratory muscles can be shown to demonstrate physiological responsesconsistent with fatigue during prolonged IRL, and activation rates arehigh and remain so throughout this prolonged loading.

     

A quantitative histological study of adrenal development during the perinatal period in intact and hypophysectomized pigs

Adrenal growth was studied between D70 of gestation (D0: day of mating) and D6 after birth in 33 fetuses and 11 new-born pigs of the Large White breed. Volume, height and mean surface, were estimated by zone, as well as cell number, cell size and nucleocytoplasmic ratio. The volume increased exponentially. In the cortex it was 4.0 mm3, 7.2 mm3, 10.5 mm3, 33.6 mm3, 55.3 mm3 at D70, D94, D106, D113 and D1-D6, respectively, after birth. In the medulla, the volume was 4.0 mm3, 6.3 mm3, 6.9 mm3, 9.6 mm3 and 20.6 mm3 at the same stages. This evolution was due to predominant longitudinal stretching between D94 and D106, then thickening, as shown by the relative variations of the height and the surface. The growth of the cortex corresponded to hyperplasia associated with hypertrophy after D110. In the medulla, hyperplasia predominated until D110; this was followed by hypertrophic phase. Twelve fetuses hypophysectomized at D44 or at D55 were recovered at D94, D106 and D112 at least 50 days after surgery. Hypophysectomy did not affect medullar development (volume and number of cells) but inhibited very significantly the growth of the cortical zone, particularly that of the fasciculata.     

Phrenic nerve conduction times and twitch pressures of the human diaphragm

A multilumen catheter was modified to allow simultaneous recording of transdiaphragmatic pressure (Pdi) and the electromyographic (EMG) activity of the diaphragm. The catheter was used in 20 healthy males to measure the conduction time of the phrenic nerves and the twitch pressure of each hemidiaphragm during single supramaximal shocks delivered to the phrenic nerve in the neck. Diaphragmatic EMG was also recorded with surface electrodes at various sites on the chest wall. The mean conduction time to the crural fibers was 6.82 +/- 0.64 ms on the right and 7.93 +/- 0.85 ms on the left, whereas that to the costal fibers adjacent to the midclavicular line was 7.68 +/- 0.56 ms on the right and 7.92 +/- 0.92 ms on the left. Significant correlations were found between the conduction time of each phrenic nerve and the height and the age of the subjects. Conduction times measured at different EMG recording sites varied by as much as 2 ms. This variability, and that of previously reported values for phrenic conduction time, may be largely accounted for by differences in the conduction distances that were measured to each site in three cadavers. The evoked change in Pdi had a mean rise time of 92 ms and an amplitude of approximately 10 cmH2O.     

An overview of phrenic nerve and diaphragm muscle development in the perinatal rat.

In this overview, we outline what is known regarding the key developmental stages of phrenic nerve and diaphragm formation in perinatal rats. These developmental events include the following. Cervical axons emerge from the spinal cord during embryonic (E) day 11. At approximately E12.5, phrenic and brachial axons from the cervical segments merge at the brachial plexi. Subsequently, the two populations diverge as phrenic axons continue to grow ventrally toward the diaphragmatic primordium and brachial axons turn laterally to grow into the limb bud. A few pioneer axons extend ahead of the majority of the phrenic axonal population and migrate along a well-defined track toward the primordial diaphragm, which they reach by E13.5. The primordial diaphragmatic muscle arises from the pleuroperitoneal fold, a triangular protrusion of the body wall composed of the fusion of the primordial pleuroperitoneal and pleuropericardial tissues. The phrenic nerve initiates branching within the diaphragm at approximately E14, when myoblasts in the region of contact with the phrenic nerve begin to fuse and form distinct primary myotubes. As the nerve migrates through the various sectors of the diaphragm, myoblasts along the nerve's path begin to fuse and form additional myotubes. The phrenic nerve intramuscular branching and concomitant diaphragmatic myotube formation continue to progress up until E17, at which time the mature pattern of innervation and muscle architecture are approximated. E17 is also the time of the commencement of inspiratory drive transmission to phrenic motoneurons (PMNs) and the arrival of phrenic afferents to the motoneuron pool. During the period spanning from E17 to birth (gestation period of approximately 21 days), there is dramatic change in PMN morphology as the dendritic branching is rearranged into the rostrocaudal bundling characteristic of mature PMNs. This period is also a time of significant changes in PMN passive membrane properties, action-potential characteristics, and firing properties.     

Maturation of the pituitary-thyroid axis during the perinatal period.

To clarify the maturation process of the pituitary-thyroid axis during the perinatal period, thyrotropin (TSH) response to thyrotropin releasing hormone (TRH) and serum thyroid hormone levels were examined in 26 healthy infants of 30 to 40 weeks gestation. A TRH stimulation test was performed on 10 to 20 postnatal days. Basal concentrations of serum thyroxine (T4), free thyroxine (free T4) and triiodothyronine (T3) were positively correlated to gestational age and birth weight (p less than 0.001-0.01). Seven infants of 30 to 35 gestational weeks demonstrated an exaggerated TSH response to TRH (49.7 +/- 6.7 microU/ml versus 22.1 +/- 4.8 microU/ml, p less than 0.001), which was gradually reduced with gestational age and normalized after 37 weeks gestation. A similar decrease in TSH responsiveness to TRH was also observed longitudinally in all of 5 high responders repeatedly examined. There was a negative correlation between basal or peak TSH concentrations and postconceptional age in high responders (r = -0.59 p less than 0.05, r = -0.66 p less than 0.01), whereas in the normal responders TSH response, remained at a constant level during 31 to 43 postconceptional weeks. On the other hand, there was no correlation between basal or peak TSH levels and serum thyroid hormones. These results indicate that (1) maturation of the pituitary-thyroid axis is intrinsically controlled by gestational age rather than by serum thyroid hormone levels, (2) hypersecretion of TSH in preterm infants induces a progressive increase in serum thyroid hormones, and (3) although there is individual variation in the maturation process, the feedback regulation of the pituitary-thyroid axis matures by approximately the 37th gestational week.     

Chromosome constitution of 500 infants dying during the perinatal period

Summary 500 chromosome results were obtained from 726 necropsied hospital perinatal deaths in London. There were 28 chromosome abnormalities (5.6% of the results). 9% of ante-partum deaths, 4% of intra-partum deaths and 6% of early neonatal deaths were chromosomally abnormal. 13% of lethally malformed infants had a chromosomal anomaly. Trisomy 18 was the most common abnormality and appears to be considerably more frequent at birth than is generally thought. Genetic disorders were found in a further 1% of perinatal deaths. The importance of the perinatal necropsy as a screening procedure for genetic disease is stressed.

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Zusammenfassung 500 Chromosomenbefunde wurden bei 726 sezierten perinatalen Todesfällen in London erhoben. Es fanden sich 28 Chromosomenanomalien (5,6%). 9% der vor der Geburt Verstorbenen, 4% der Verstorbenen unter der Geburt sowie 6% der Todesfälle lurz danach zeigten eine Chromosomenanomalie. 13% der letal mißgebildeten Kinder wiesen einen Chromosomenbefund auf. Trisomie 18 was am häufigsten; sie scheint zum Zeitpunkt der Geburt wesentlich häufiger zu sein, als bisher angenommen. Bei einem weiteren Prozent der verstorbenen Neugeborenen wurden genetisch bedingte Erkrankungen entdeckt. Es wird die Bedeutung der Sektion perinataler Todesfälle als Screeningverfahren für genetisch bedingte Erkrankungen betont.

     

Osteocalcin is vitamin D-dependent during the perinatal period in the rat

The vitamin D-dependence of renal calbindin D-28K and osteocalcin during the perinatal period was studied in fetuses (days 18 and 21) and neonates (days 2, 12, 17 and 22) of rats fed either a standard diet (0.85% Ca-0.7% P; "high Ca-P diet" rats) or a mildly Ca-P restricted diet (0.2% Ca-0.2% P; "low Ca-P diet" rats). Body weight and plasma calcium levels were identical in both groups. Plasma 1,25(OH)2D concentrations were markedly higher in the low Ca-P diet rats at all stages of fetal and neonatal life (in 22-day-old neonates: 536 +/- 58 pg/ml versus 126 +/- 12 pg/ml). 1,25(OH)2D concentrations increased between day 18 and 21 of fetal life, remained constant between day 21 of fetal and day 12 of neonatal life, and increased sharply between day 12 and 17 in both groups; after day 17, 1,25(OH)2D concentrations increased further in pups fed the low Ca-P diet. Renal calbindin D-28K reached peak concentrations on day 12 of neonatal life; calbindin D-28K levels were similar in the high and low Ca-P diet rats at all stages of perinatal development. Plasma osteocalcin levels increased steadily during the perinatal period; at most stages of perinatal life, and already from the fetal period was osteocalcin higher in the low Ca-P diet rats than in the high Ca-P diet rats (in 22-day-old pups: 1106 +/- 47 ng/ml versus 429 +/- 14 ng/ml). Femoral osteocalcin concentrations were also increased in fetal and early neonatal (days 2 and 12) low Ca-P diet rats, while the femoral calcium content and concentration of these rats were decreased in the late neonatal period (days 12, 17 and 22). These studies indicate that osteocalcin is vitamin D-dependent in the fetal and neonatal rat.     

围产期无乳链球菌携带状况及耐药性分析

目的 探讨新疆乌鲁木齐市围产期无乳链球菌携带状况及耐药性,为临床合理使用抗菌药物提供依据。方法 回顾性分析2014年1月—2016年11月围产期6 003例泌尿生殖道分泌物标本,采用法国梅里埃Vitek 2 Compact全自动细菌鉴定和药敏系统检测,从中检出151株无乳链球菌,分析药敏、查阅其病历基本信息、新生儿出生状况等。结果 6 003例围产期泌尿生殖道标本中检出151株无乳链球菌,检出率为2.52%,其中≤35岁、>35岁孕产妇中无乳链球菌感染率分别为2.55%、2.26%,两者相比差异无统计学意义（P>0.05）;民族中,汉族、维吾尔族、回族、哈萨克族中无乳链球菌感染率分别为2.70%、2.63%、2.18%、1.73%,相比差异无统计学意义（P>0.05）;>37周感染无乳链球菌101例且诊断胎膜早破有76例;孕1次感染无乳链球菌有90例,孕3次感染无乳链球菌有33例,且有20例曾有流产史;新生儿宫内感染中有1例感染无乳链球菌属于早发型。药敏结果中,无乳链球菌对氨苄西林、青霉素G敏感率分别为97.97%、98.63%;对红霉素、四环素、克林霉素和左氧氟沙星耐药率较高,分别为77.46%、72.85%、58.94%和54.30%。结论 临床应重视在围产期对无乳链球菌的检测和耐药情况,合理使用抗生素,可有效地预防和治疗该菌的感染。     

Hypophyseal corticotropic activity during perinatal period: ontogenesis and regulation

1. In the fetal rat hypophysis, the "opio-melano-corticotropic" cells were immunocytologically revealed earlier in the pars distalis (day 16) than in the pars intermedia (day 17). At an early stage of embryonic development, some factors, possibly of nervous origin, could exert inductive influence on the differentiation and/or the increase in the number of immunoreactive cells. 2. Several forms of ACTH, characterized by their apparent molecular weight, bioactivity and immunoreactivity, can be observed when acid extracts are subjected to gel filtration on Sephadex G 50 fine columns. The ratios between these ACTH forms change during fetal development; the rise in bioactive ACTH content during the last days of gestation, is correlated with an increase of the "intermediate" and "little" molecular forms of ACTH and a parallel decrease of the "big" form. 3. The hypophyseal corticotropic activity is highest on days 18-19 of gestation. Between days 17 and 19, the cortico-stimulating activity is largely independent of the hypothalamus although the fetal hypothalamus shows light hypophyso-stimulating influence. In contrast, between days 19 and 21, the hypophyseal corticotropic activity is mostly under the hypothalamic control; the ACTH release is greatly reduced in its absence. 4. Circulating corticosteroids exert a negative feedback directly at the hypothalamic and pituitary levels. Such negative feedback could explain the decrease of the corticotropic activity of the pituitary during the last three days of gestation and the first postnatal days. 5. The fetus at the end of gestation, as the newborn, is responsive to stress; however, the extent of the pituitary response is weaker than in the adult.