Pharmacological Interventions in Labor and Delivery.

While there is not a wide range of pregnancy-specific drugs, there are some very specific high-risk areas of obstetric care for which unique pharmacological approaches have been established. In preterm birth, labor induction and augmentation, and the management of postpartum hemorrhage, these pharmacological approaches have become the bedrock in managing some of the most common and problematic areas of antenatal and intrapartum care. In this review, we summarize the existing established and emerging evidence that supports and broadens these pharmacological approaches to obstetric management and its impact on clinical practice. It is clear that existing therapeutics are limited. They have largely been developed from our knowledge of the physiology of the myometrium and act on hormonal receptors and their signaling pathways or on ion channels influencing excitability. Newer drugs in development are mostly refinements of these two approaches, but novel agents from plants and improved formulations are also discussed.

Uterine Excitability and Ion Channels and Their Changes with Gestation and Hormonal Environment.

We address advances in the understanding of myometrial physiology, focusing on excitation and the effects of gestation on ion channels and their relevance to labor. This review moves through pioneering studies to exciting new findings. We begin with the myometrium and its myocytes and describe how excitation might initiate and spread in this myogenic smooth muscle. We then review each of the ion channels in the myometrium: L- and T-type Ca2+ channels, KATP (Kir6) channels, voltage-dependent K channels (Kv4, Kv7, and Kv11), twin-pore domain K channels (TASK, TREK), inward rectifier Kir7.1, Ca2+-activated K+ channels with large (KCNMA1, Slo1), small (KCNN1-3), and intermediate (KCNN4) conductance, Na-activated K channels (Slo2), voltage-gated (SCN) Na+ and Na+ leak channels, nonselective (NALCN) channels, the Na K-ATPase, and hyperpolarization-activated cation channels. We finish by assessing how three key hormones- oxytocin, estrogen, and progesterone-modulate and integrate excitability throughout gestation.

Drebrin regulates cytoskeleton dynamics in migrating neurons through interaction with CXCR4.

Stromal cell-derived factor-1 (SDF-1) and chemokine receptor type 4 (CXCR4) are regulators of neuronal migration (e.g., GnRH neurons, cortical neurons, and hippocampal granule cells). However, how SDF-1/CXCR4 alters cytoskeletal components remains unclear. Developmentally regulated brain protein (drebrin) stabilizes actin polymerization, interacts with microtubule plus ends, and has been proposed to directly interact with CXCR4 in T cells. The current study examined, in mice, whether CXCR4 under SDF-1 stimulation interacts with drebrin to facilitate neuronal migration. Bioinformatic prediction of protein-protein interaction highlighted binding sites between drebrin and crystallized CXCR4. In migrating GnRH neurons, drebrin, CXCR4, and the microtubule plus-end binding protein EB1 were localized close to the cell membrane. Coimmunoprecipitation (co-IP) confirmed a direct interaction between drebrin and CXCR4 using wild-type E14.5 whole head and a GnRH cell line. Analysis of drebrin knockout (DBN1 KO) mice showed delayed migration of GnRH cells into the brain. A decrease in hippocampal granule cells was also detected, and co-IP confirmed a direct interaction between drebrin and CXCR4 in PN4 hippocampi. Migration assays on primary neurons established that inhibiting drebrin (either pharmacologically or using cells from DBN1 KO mice) prevented the effects of SDF-1 on neuronal movement. Bioinformatic prediction then identified binding sites between drebrin and the microtubule plus end protein, EB1, and super-resolution microscopy revealed decreased EB1 and drebrin coexpression after drebrin inhibition. Together, these data show a mechanism by which a chemokine, via a membrane receptor, communicates with the intracellular cytoskeleton in migrating neurons during central nervous system development.

Nitric oxide resets kisspeptin-excited GnRH neurons via PIP2 replenishment.

Fertility relies upon pulsatile release of gonadotropin-releasing hormone (GnRH) that drives pulsatile luteinizing hormone secretion. Kisspeptin (KP) neurons in the arcuate nucleus are at the center of the GnRH pulse generation and the steroid feedback control of GnRH secretion. However, KP evokes a long-lasting response in GnRH neurons that is hard to reconcile with periodic GnRH activity required to drive GnRH pulses. Using calcium imaging, we show that 1) the tetrodotoxin-insensitive calcium response evoked by KP relies upon the ongoing activity of canonical transient receptor potential channels maintaining voltage-gated calcium channels in an activated state, 2) the duration of the calcium response is determined by the rate of resynthesis of phosphatidylinositol 4,5-bisphosphate (PIP2), and 3) nitric oxide terminates the calcium response by facilitating the resynthesis of PIP2 via the canonical pathway guanylyl cyclase/3',5'-cyclic guanosine monophosphate/protein kinase G. In addition, our data indicate that exposure to nitric oxide after KP facilitates the calcium response to a subsequent KP application. This effect was replicated using electrophysiology on GnRH neurons in acute brain slices. The interplay between KP and nitric oxide signaling provides a mechanism for modulation of the refractory period of GnRH neurons after KP exposure and places nitric oxide as an important component for tonic GnRH neuronal pulses.

Novel factor in olfactory ensheathing cell-astrocyte crosstalk: Anti-inflammatory protein α-crystallin B.

Astrocytes are key players in CNS neuroinflammation and neuroregeneration that may help or hinder recovery, depending on the context of the injury. Although pro-inflammatory factors that promote astrocyte-mediated neurotoxicity have been shown to be secreted by reactive microglia, anti-inflammatory factors that suppress astrocyte activation are not well-characterized. Olfactory ensheathing cells (OECs), glial cells that wrap axons of olfactory sensory neurons, have been shown to moderate astrocyte reactivity, creating an environment conducive to regeneration. Similarly, astrocytes cultured in medium conditioned by cultured OECs (OEC-CM) show reduced nuclear translocation of nuclear factor kappa-B (NFκB), a pro-inflammatory protein that induces neurotoxic reactivity in astrocytes. In this study, we screened primary and immortalized OEC lines to identify these factors and discovered that Alpha B-crystallin (CryAB), an anti-inflammatory protein, is secreted by OECs via exosomes, coordinating an intercellular immune response. Our results showed that: (a) OEC exosomes block nuclear NFκB translocation in astrocytes while exosomes from CryAB-null OECs could not; (b) OEC exosomes could be taken up by astrocytes, and (c) CryAB treatment suppressed neurotoxicity-associated astrocyte transcripts. Our results indicate CryAB, as well as other factors secreted by OECs, are potential agents that can ameliorate, or even reverse, the growth-inhibitory environment created by neurotoxic reactive astrocytes following CNS injuries.

HYPOXIA AND REPRODUCTIVE HEALTH: Hypoxia and labour.

Intermittent myometrial hypoxia is a normal feature of labour, as the powerful contractions compress blood vessels. In this review, we focus on the relation between hypoxia, myometrial metabolism, and contractility. We dissect how hypoxia can feedback and limit an ongoing contraction and help prevent foetal distress. The mechanisms involve acidification from lactate, decreased excitability, and a fall of intracellular calcium concentration. As this cycle of contraction and relaxation repeats in labour, the hypoxia also engenders mechanisms that increase force; hypoxia-induced force increase, HIFI. We also discuss the role of the myometrial blood vessels in dysfunctional labour, which is associated with lactic acidosis. In synthesising these studies, we have attempted to unify findings by considering the importance of experimental protocols and finding direct mechanistic evidence from human myometrium or in vivo studies. We have made suggestions for future studies to fill the holes in our understanding and speed up the translation of our knowledge to improve births for mothers and babies everywhere.

Do stress and anxiety in early pregnancy affect the progress of labor: Evidence from the Wirral Child Health and Development Study.

INTRODUCTION: Despite widespread belief that anxiety causes longer labor, evidence of association is inconsistent. Data gathered as part of a prospective epidemiological longitudinal study were used to investigate associations between antenatal anxiety and pregnancy-specific stress, and labor progression was assessed by duration and use of augmentation. MATERIAL AND METHODS: Pregnant primiparous women completed measures for anxiety and pregnancy-specific stress at 20 weeks' gestation (n = 1145). Birth outcome data were extracted from medical records. Regression analyses and a path analysis assessed associations between antenatal anxiety and pregnancy-specific stress, and indices of labor progression (labor duration and augmentation). RESULTS: Anxiety/pregnancy-specific stress were not directly associated with duration of stage 1 labor (HIGH/LOW anxiety: mean difference = 13.94 minutes, SD = 20.66, 95% CI -26.60 to 54.49, P < .50)/(HIGH/LOW pregnancy-specific stress: mean difference = 12.05 minutes, SD = 16.09, 95% CI -19.52 to 43.63, P < .45). However, anxiety/pregnancy-specific stress were associated with epidural use (HIGH/LOW anxiety: 39% vs 31%, P < .042; HIGH/LOW pregnancy-specific stress: 38% vs 29%, P < .001), which was itself associated with longer labor (mean difference: 158.79 minutes, SD = 16.76, 95% CI 125.89-191.68, P < .001). Anxiety and pregnancy-specific stress were associated with increased likelihood of augmentation but these associations were nonsignificant after accounting for epidural, which was itself highly associated with augmentation. However, path analysis indicated an indirect effect linking pregnancy-specific stress, but not general anxiety, to labor duration and augmentation: elevated pregnancy-specific stress led to greater use of epidural, which was linked to both increased rates of augmentation, and increased labor duration. CONCLUSIONS: Contrary to general belief, general anxiety and specific pregnancy stress were not directly linked to longer duration of stage one labor. However specific pregnancy stress was associated with epidural use, which in turn was significantly associated with risk of augmentation, and longer stage one labor. Identification of pregnancy-specific stress could help to identify women for whom psychological interventions could improve birth experience.

Justicia flava leaf extract potently relaxes pregnant human myometrial contractility: a lead plant for drug discovery of new tocolytic drugs.

NEW FINDINGS: What is the central question of this study? Can Justicia flava leaf extract (JF) inhibit human myometrial contractility as was previously shown in mouse myometrium? What is the main finding and its importance? JF abolished human myometrial contractions and therefore presents as a lead plant in drug discovery studies involving drugs for preterm birth. ABSTRACT: In the search for new potent therapies for preterm labour, Justicia flava leaf extract (JF) was previously shown to potently inhibit uterine contractility in both pregnant and non-pregnant mouse uterus. This study took the investigation a step further and investigated the activity of JF on pregnant human myometrial contractility. JF potently inhibited human myometrial contractility in a concentration-dependent manner. This pilot study provides evidence that JF should be further investigated as a lead plant in the drug discovery of new uterine relaxants.

The Myometrium: From Excitation to Contractions and Labour.

We start by describing the functions of the uterus, its structure, both gross and fine, innervation and blood supply. It is interesting to note the diversity of the female's reproductive tract between species and to remember it when working with different animal models. Myocytes are the overwhelming cell type of the uterus (>95%) and our focus. Their function is to contract, and they have an intrinsic pacemaker and rhythmicity, which is modified by hormones, stretch, paracrine factors and the extracellular environment. We discuss evidence or not for pacemaker cells in the uterus. We also describe the sarcoplasmic reticulum (SR) in some detail, as it is relevant to calcium signalling and excitability. Ion channels, including store-operated ones, their contributions to excitability and action potentials, are covered. The main pathway to excitation is from depolarisation opening voltage-gated Ca2+ channels. Much of what happens downstream of excitability is common to other smooth muscles, with force depending upon the balance of myosin light kinase and phosphatase. Mechanisms of maintaining Ca2+ balance within the myocytes are discussed. Metabolism, and how it is intertwined with activity, blood flow and pH, is covered. Growth of the myometrium and changes in contractile proteins with pregnancy and parturition are also detailed. We finish with a description of uterine activity and why it is important, covering progression to labour as well as preterm and dysfunctional labours. We conclude by highlighting progress made and where further efforts are required.

Hypoxic conditioning in blood vessels and smooth muscle tissues: effects on function, mechanisms, and unknowns.

Hypoxic preconditioning, the protective effect of brief, intermittent hypoxic or ischemic episodes on subsequent more severe hypoxic episodes, has been known for 30 yr from studies on cardiac muscle. The concept of hypoxic preconditioning has expanded; excitingly, organs beyond the heart, including the brain, liver, and kidney, also benefit. Preconditioning of vascular and visceral smooth muscles has received less attention despite their obvious importance to health. In addition, there has been no attempt to synthesize the literature in this field. Therefore, in addition to overviewing the current understanding of hypoxic conditioning, in the present review, we consider the role of blood vessels in conditioning and explore evidence for conditioning in other smooth muscles. Where possible, we have distinguished effects on myocytes from other cell types in the visceral organs. We found evidence of a pivotal role for blood vessels in conditioning and for conditioning in other smooth muscle, including the bladder, vascular myocytes, and gastrointestinal tract, and a novel response in the uterus of a hypoxic-induced force increase, which helps maintain contractions during labor. To date, however, there are insufficient data to provide a comprehensive or unifying mechanism for smooth muscles or visceral organs and the effects of conditioning on their function. This also means that no firm conclusions can be drawn as to how differences between smooth muscles in metabolic and contractile activity may contribute to conditioning. Therefore, we have suggested what may be general mechanisms of conditioning occurring in all smooth muscles and tabulated tissue-specific mechanistic findings and suggested ideas for further progress.

Hypoxia-induced force increase (HIFI) is a novel mechanism underlying the strengthening of labor contractions, produced by hypoxic stresses.

For successful birth, contractions need to become progressively stronger. The underlying mechanisms are unknown, however. We have found that a novel mechanism, hypoxia-induced force increase (HIFI), is switched on selectively, at term, and is essential to strengthening contractions. HIFI is initiated as contractions cyclically reduce blood flow and produce repeated hypoxic stresses, with associated metabolic and transcriptomic changes. The increases in contractility are a long-lasting, oxytocin-independent, intrinsic mechanism present only in the full-term pregnant uterus. HIFI is inhibited by adenosine receptor antagonism and blockade of cyclooxygenase-2 signaling, and partially reproduced by brief episodes of acidic (but not alkalotic) pH. HIFI explains how labor can progress despite paradoxical metabolic challenge, and provides a new mechanistic target for the 1 in 10 women suffering dysfunctional labor because of poor contractions.

Sarcoplasmic reticulum function in smooth muscle.

The sarcoplasmic reticulum (SR) of smooth muscles presents many intriguing facets and questions concerning its roles, especially as these change with development, disease, and modulation of physiological activity. The SR's function was originally perceived to be synthetic and then that of a Ca store for the contractile proteins, acting as a Ca amplification mechanism as it does in striated muscles. Gradually, as investigators have struggled to find a convincing role for Ca-induced Ca release in many smooth muscles, a role in controlling excitability has emerged. This is the Ca spark/spontaneous transient outward current coupling mechanism which reduces excitability and limits contraction. Release of SR Ca occurs in response to inositol 1,4,5-trisphosphate, Ca, and nicotinic acid adenine dinucleotide phosphate, and depletion of SR Ca can initiate Ca entry, the mechanism of which is being investigated but seems to involve Stim and Orai as found in nonexcitable cells. The contribution of the elemental Ca signals from the SR, sparks and puffs, to global Ca signals, i.e., Ca waves and oscillations, is becoming clearer but is far from established. The dynamics of SR Ca release and uptake mechanisms are reviewed along with the control of luminal Ca. We review the growing list of the SR's functions that still includes Ca storage, contraction, and relaxation but has been expanded to encompass Ca homeostasis, generating local and global Ca signals, and contributing to cellular microdomains and signaling in other organelles, including mitochondria, lysosomes, and the nucleus. For an integrated approach, a review of aspects of the SR in health and disease and during development and aging are also included. While the sheer versatility of smooth muscle makes it foolish to have a "one model fits all" approach to this subject, we have tried to synthesize conclusions wherever possible.

Mutations in FEZF1 cause Kallmann syndrome.

Gonadotropin-releasing hormone (GnRH) neurons originate outside the CNS in the olfactory placode and migrate into the CNS, where they become integral components of the hypothalamic-pituitary-gonadal (HPG) axis. Disruption of this migration results in Kallmann syndrome (KS), which is characterized by anosmia and pubertal failure due to hypogonadotropic hypogonadism. Using candidate-gene screening, autozygosity mapping, and whole-exome sequencing in a cohort of 30 individuals with KS, we searched for genes newly associated with KS. We identified homozygous loss-of-function mutations in FEZF1 in two independent consanguineous families each with two affected siblings. The FEZF1 product is known to enable axons of olfactory receptor neurons (ORNs) to penetrate the CNS basal lamina in mice. Because a subset of axons in these tracks is the migratory pathway for GnRH neurons, in FEZF1 deficiency, GnRH neurons also fail to enter the brain. These results indicate that FEZF1 is required for establishment of the central component of the HPG axis in humans.

GnRH, anosmia and hypogonadotropic hypogonadism--where are we?

Gonadotropin releasing hormone (GnRH) neurons originate the nasal placode and migrate into the brain during prenatal development. Once within the brain, these cells become integral components of the hypothalamic-pituitary-gonadal axis, essential for reproductive function. Disruption of this system causes hypogonadotropic hypogonadism (HH). HH associated with anosmia is clinically defined as Kallman syndrome (KS). Recent work examining the developing nasal region has shed new light on cellular composition, cell interactions and molecular cues responsible for the development of this system in different species. This review discusses some developmental aspects, animal models and current advancements in our understanding of pathologies affecting GnRH. In addition we discuss how development of neural crest derivatives such as the glia of the olfactory system and craniofacial structures control GnRH development and reproductive function.

The combination tocolytic effect of magnesium sulfate and an oxytocin receptor antagonist in myometrium from singleton and twin pregnancies.

BACKGROUND: Preterm birth at <37 weeks of gestation is the most common and costly complication of pregnancy and remains the leading cause of neonatal morbidity, death, and reduced achievement in surviving infants. Magnesium sulfate is 1 class of tocolytics for threatened preterm labor; however, its clinical efficacy has been questioned. Twin pregnancies are at increased risk of preterm delivery compared with singleton gestations, which suggests that there is twin-specific risk to preterm delivery in twins. The prevention strategies that are applied to singleton pregnancies, however, have not been shown to be effective in twin pregnancies. OBJECTIVE: The purpose of this study was to compare the relaxant effect of magnesium sulfate on spontaneous and oxytocin-augmented contractions of human myometrium from singleton and twin pregnancies and to examine whether the effect of oxytocin on magnesium sulfate's potency could be reversed with the use of the oxytocin receptor antagonist, atosiban. STUDY DESIGN: Myometrium was obtained at the time of prelabor cesarean section (36-40 weeks of gestation) from women with singleton (n=23) or twin (n=12) pregnancy. Isometric tension recordings were made on myometrial strips that were mounted in organ baths that were superfused with physiologic saline solution. Strips were exposed to rising concentrations of magnesium sulfate, and the effect on spontaneous contractions or stimulated with oxytocin (0.5 nmol/L) and in the presence or absence of atosiban (100 nmol/L) was recorded. The contractile characteristics after each application of magnesium sulfate, which included amplitude of contraction and activity integral, were measured. Concentration-response curves were fitted with the use of nonlinear regression and comparison of the negative logarithm of the 50% reduction in activity values. RESULTS: Magnesium sulfate exerted an equal concentration-dependent inhibitory effect on spontaneous myometrial contractions from both singleton and twin myometrium (P>.05). The application of oxytocin produced a significant rightward shift in the concentration-response curves (P<.0001), but no differences were found between pregnancy groups (P>.05). The addition of atosiban shifted concentration-response curves significantly back to the left for amplitude of contraction and activity integral in singletons (P<.0001). However, only activity integral was significantly reversed in twins (P<.01). CONCLUSION: Magnesium sulfate is equipotent in suppressing contractions in singleton and twin myometrium. Oxytocin (0.5 nmol/L) significantly reduces the tocolytic potency of magnesium sulfate, which may explain, in part, magnesium sulfate's poor efficacy in vivo; however, this can be reversed partially by the use of an oxytocin receptor antagonist. Combination tocolysis that involves oxytocin receptor antagonists requires further investigation.

Oxytocic plant cyclotides as templates for peptide G protein-coupled receptor ligand design.

Cyclotides are plant peptides comprising a circular backbone and three conserved disulfide bonds that confer them with exceptional stability. They were originally discovered in Oldenlandia affinis based on their use in traditional African medicine to accelerate labor. Recently, cyclotides have been identified in numerous plant species of the coffee, violet, cucurbit, pea, potato, and grass families. Their unique structural topology, high stability, and tolerance to sequence variation make them promising templates for the development of peptide-based pharmaceuticals. However, the mechanisms underlying their biological activities remain largely unknown; specifically, a receptor for a native cyclotide has not been reported hitherto. Using bioactivity-guided fractionation of an herbal peptide extract known to indigenous healers as "kalata-kalata," the cyclotide kalata B7 was found to induce strong contractility on human uterine smooth muscle cells. Radioligand displacement and second messenger-based reporter assays confirmed the oxytocin and vasopressin V1a receptors, members of the G protein-coupled receptor family, as molecular targets for this cyclotide. Furthermore, we show that cyclotides can serve as templates for the design of selective G protein-coupled receptor ligands by generating an oxytocin-like peptide with nanomolar affinity. This nonapeptide elicited dose-dependent contractions on human myometrium. These observations provide a proof of concept for the development of cyclotide-based peptide ligands.

Physiological increases in lactate inhibit intracellular calcium transients, acidify myocytes and decrease force in term pregnant rat myometrium.

Lactate is increased in myometrial capillary blood from women in slow or non-progressive labour (dystocia), suggesting that it is detrimental to uterine contractions. There are, however, no studies of the effect of lactate on the myometrium. We therefore investigated its effects and mechanism of action on myometrial strips from term pregnant rats. The effects on spontaneous and oxytocin-induced contractility in response to sodium lactate and other weak acids (1-20 mM) were studied. In some experiments, simultaneous force and intracellular Ca(2+) or pH (pH(i)) were measured with Indo-1 or Carboxy-SNARF, respectively. Statistical differences were tested using non-parametric tests. Lactate significantly decreased spontaneous contractility with an EC50 of 3.9 mM. Propionate, butyrate and pyruvate also reduced contractions with similar potency. The effects of lactate were reduced in the presence of oxytocin but remained significant. Lactate decreased pH(i) and nulling the decrease in pH(i) abolished its effects. We also show that lactate inhibited Ca(2+) transients, with these changes mirroring those produced on force. If Ca(2+) entry was enhanced by depolarization (high KCl) or applying the Ca(2+) channel agonist, Bay K 4644, the effects of lactate were abolished. Taken together, these data show that lactate in the physiological range potently decreases myometrial contractility as a result of its inhibition of Ca(2+) transients, which can be attributed to the induced acidification. The present study suggests that the accumulation of extracellular lactate will reduce myometrial contractions and could therefore contribute to labour dystocia.

Insights from physiology into myometrial function and dysfunction.

NEW FINDINGS: What is the topic of this review? I focus on clinical aspects of uterine physiology, specifically, myometrial contractility. I bring together and contrast findings using physiological approaches and those using newer techniques, 'omics'. What advances does it highlight? Physiological studies have recently shed light on the myometrium in twin pregnancies, but there have been no 'omic' approaches. In contrast, studies of preterm delivery using newer approaches are generating new research avenues, whereas traditional approaches have not flourished. Finally, I describe significant advances in understanding of 'slow-to-progress' labours, achieved using physiological and clinical approaches. Advances in molecular, genetic and 'omic' technologies are fuelling the thirst for better understanding of the uterus and application of this information to problems in pregnancy and labour. Progress has, however, been limited while we still have an incomplete understanding of some of the basic physiology of uterine smooth muscle (myometrium). In this review and opinion piece, I explore some of the fascinating findings from selected recent studies and see how these may provide new avenues for physiological and clinical research. It is also the case, however, that there is still limited mechanistic understanding about physiological and pathophysiological processes in the myometrium. This lack of understanding limits the usefulness of some findings from genomic and allied studies. By focusing on some key recent findings and relating these to two important clinical problems in childbirth that involve myometrial activity, namely preterm delivery and difficult labours, the interplay between our physiological knowledge and the information provided by newer technologies is explored. My opinion is that physiology has provided much more new mechanistic insight into difficult births and that the newer technologies may lead to breakthroughs in preterm birth research, but that this has not yet happened.

Calcium release-dependent actin flow in the leading process mediates axophilic migration.

Proper assembly of neural circuits requires newly born neurons to migrate from their place of origin to their final location. Little is known about the mechanisms of axophilic neuronal migration, whereby neurons travel along axon pathways to navigate to their destinations. Gonadotropin-releasing hormone (GnRH)-expressing neurons migrate along olfactory axons from the nose into the forebrain during development, and were used as a model of axophilic migration. After migrating, GnRH neurons are located in the hypothalamus and are essential for puberty and maintenance of reproductive function. To gain a better understanding of the mechanisms underlying axophilic migration, we investigated in mice the regulation of movement from calcium signals to cytoskeletal dynamics. Live imaging revealed robust calcium activity during axophilic migration, and calcium release through IP3 receptors was found to stimulate migration. This occurred through a signaling pathway involving the calcium sensor calcium/calmodulin protein kinase kinase, AMP-activated kinase, and RhoA/ROCK. By imaging GnRH neurons expressing actin-GFP or Lifeact-RFP, calcium release was found to stimulate leading process actin flow away from the cell body. In contrast, actin contractions at the cell rear were unaffected by this calcium signaling pathway. These findings are the first to test the regulation of cytoskeletal dynamics in axophilic migration, and reveal mechanisms of movement that have broad implications for the migration of other CNS populations.