Expression and androgen regulation of the ciliary neurotrophic factor receptor (CNTFRα) in muscles and spinal cord

We have previously observed that ciliary neurotrophic factor (CNTF) can prevent the degeneration of androgen-sensitive perineal motoneurons and their target muscles, the bulbocavernosus and levator ani (BC/LA), in perinatal female rats. Response to CNTF is dependent on the expression of the alpha component of the CNTF receptor (CNTFRα). In the present study, we examined the developmental profile and androgen regulation of CNTFRα gene expression in BC/LA muscle, thigh muscle, and lumbosacral spinal cord. CNTFRα mRNA was abundantly expressed in the BC/LA and thigh around the time of birth; expression declined progressively after birth and remained low into adulthood. In contrast, CNTFRα message remained high in the lumbosacral spinal cord throughout development. Androgen regulation of CNTFRα expression was examined in prenatal animals by administering the androgen receptor blocker hydroxyflutamide from embryonic days E18 through E21. Four days of androgen deprivation caused a significant up-regulation of CNTFRα mRNA in the BC/LA, thigh, and spinal cord of male fetuses. After castration in adulthood, CNTFRα expression in the BC/LA transiently increased, then decreased below control levels. Expression of CNTFRα in thigh muscles and the lumbosacral spinal cord was not affected by adult castration. Thus, the perineal muscles and motoneurons are potential sites of direct CNTF action, and expression of the CNTFRα gene is modulated by androgen, especially in the androgen-sensitive perineal muscles. Transient up-regulation of CNTFRα following castration or androgen receptor blockade may represent a protective response designed to counteract the muscle atrophy normally induced by androgen withdrawal. © 1998 John Wiley & Sons, Inc. J Neurobiol 35: 217–225, 1998     

A phylogenetic approach to the identification of phosphoglucomutase genes

The expanding molecular database provides unparalleled opportunities for characterizing genes and for studying groups of related genes. We use sequences drawn from the database to construct an evolutionary framework for examining the important glycolytic enzyme phosphoglucomutase (PGM). Phosphoglucomutase plays a pivotal role in the synthesis and utilization of glycogen and is present in all organisms. In humans, there are three well-described isozymes, PGMI, PGM2, and PGM3. PGM1 was cloned 5 years ago; however, repeated attempts using both immunological approaches and molecular probes designed from PGM1 have failed to isolate either PGM2 or PGM3. Using a phylogenetic strategy, we first identified 47 highly divergent prokaryotic and eukaryotic PGM-like sequences from the database. Although overall amino acid identity often fell below 20%, the relative order, position, and sequence of three structural motifs, the active site and the magnesium-- and sugar-binding sites, were conserved in all 47 sequences. The phylogenetic history of these sequences was complex and marked by duplications and translocations; two instances of transkingdom horizontal gene transfer were identified. Nonetheless, the sequences fell within six well-defined evolutionary lineages, three of which contained only prokaryotes. Of the two prokaryotic/eukaryotic lineages, one contained bacterial, yeast, slimemold, invertebrate, and vertebrate homologs to human PGM1 and the second contained likely homologs to human PGM2. Indeed, an amino acid sequence, derived from a partial human cDNA, that fell within the second cross-kingdom lineage bears several characteristics expected for PGM2. A third lineage may contain homologs to human PGM3. On a general level, our phylogenetic-based approach shows promise for the further utilization of the extensive molecular database.      

Incorporation of axonally transported glycoproteins into axolemma during nerve regeneration

The insertion of axonally transported fucosyl glycoproteins into the axolemma of regenerating nerve sprouts was examined in rat sciatic motor axons at intervals after nerve crush. [(3)H]Fucose was injected into the lumbar ventral horns and the nerves were removed at intervals between 1 and 14 d after labeling. To follow the fate of the “pulse- labeled” glycoproteins, we examined the nerves by correlative radiometric and EM radioautographic approaches. The results showed, first, that rapidly transported [(3)H]fucosyl glycoproteins were inserted into the axolemma of regenerating sprouts as well as parent axons. At 1 d after delivery, in addition to the substantial mobile fraction of radioactivity still undergoing bidirectional transport within the axon, a fraction of label was already associated with the axolemma. Insertion of labeled glycoproteins into the sprout axolemma appeared to occur all along the length of the regenerating sprouts, not just in sprout terminals. Once inserted, labeled glycoproteins did not undergo extensive redistribution, nor did they appear in sprout regions that formed (as a result of continued outgrowth) after their insertion. The amount of radioactivity in the regenerating nerves decreased with time, in part as a result of removal of transported label by retrograde transport. By 7-14 d after labeling, radioautography showed that almost all the remaining radioactivity was associated with axolemma. The regenerating sprouts retained increased amounts of labeled glycoproteins; 7 or 14 d after labeling, the regenerating sprouts had over twice as much of radioactivity as comparable lengths of control nerves or parent axons. One role of fast axonal transport in nerve regeneration is the contribution to the regenerating sprout of glycoproteins inserted into the axolemma; these membrane elements are added both during longitudinal outgrowth and during lateral growth and maturation of the sprout.     

Statistical properties of the variation at linked microsatellite loci: implications for the history of human Y chromosomes [published erratum appears in Mol Biol Evol 1997 Mar;14(3):354]

It has recently been suggested that observed levels of variation at microsatellite loci can be used to infer patterns of selection in genomes and to assess demographic history. In order to evaluate the feasibility of these suggestions it is necessary to know something about how levels of variation at microsatellite loci are expected to fluctuate due simply to stochasticity in the processes of mutation and inheritance (genetic sampling). Here we use recently derived properties of the stepwise mutation model to place confidence intervals around the variance in repeat score that is expected at mutation-drift equilibrium and outline a statistical test for whether an observed value differs significantly from expectation. We also develop confidence intervals for the time course of the buildup of variation following a complete elimination of variation, such as might be caused by a selective sweep or an extreme population bottleneck. We apply these methods to the variation observed at human Y-specific microsatellites. Although a number of authors have suggested the possibility of a very recent sweep, our analyses suggest that a sweep or extreme bottleneck is unlikely to have occurred anytime during the last approximately 74,000 years. To generate this result we use a recently estimated mutation rate for microsatellite loci of 5.6 x 10(-4) along with the variation observed at autosomal microsatellite loci to estimate the human effective population size. This estimate is 18,000, implying an effective number of 4,500 Y chromosomes. One important general conclusion to emerge from this study is that in order to reject mutation-drift equilibrium at a set of linked microsatellite loci it is necessary to have an unreasonably large number of loci unless the observed variance is far below that expected at mutation-drift equilibrium.      

Influence of gonadal sex hormones on behavioral components of the reproductive hierarchy in naked mole-rats

Naked mole-rats (Heterocephalus glaber) are fossorial, eusocial rodents that live in colonies which typically include 60-80 individuals. Generally, only one of the females and 1-3 of the males in a colony are reproductives. The reproductives engage in mutual genital nuzzling behavior that is rarely exhibited by subordinates (non-reproductives). Thus, genital nuzzling may represent a mechanism of bonding and/or specific recognition between reproductive individuals. We investigated whether gonadal hormones are involved in the maintenance of genital nuzzling behavior and mating behaviors in isolated pairs of mole-rats and also in established breeding pairs of mole-rats within colonies. We also explored whether sex hormone deprivation would alter the strict partner preference for performance of nuzzling within colonies. Our results indicate (a) considerable variation between pairs in the frequency of nuzzling, (b) a reduction in the frequency of nuzzling following castration of the male and restoration of the 'baseline' frequency after replacement of testosterone in castrated males, (c) the failure of either castration or combined castration and ovariectomy to eliminate genital nuzzling in established pairs, and (d) the exhibition of nuzzling behavior by some of the subordinates in all three experimental colonies beginning several weeks after gonadectomy of both of the reproductives. No cases of lordosis behavior were seen during the approximately 109 h of behavioral observations. This is not surprising, since female mole-rats have an approximately 30-day ovulatory cycle, and lordosis only occurs during a peri-ovulatory period of a few hours. A total of 44 cases of mounting behavior were recorded; all these involved breeding males in colonies or males from isolated pairs, and all occurred when males were either gonad-intact or castrated with testosterone replacement. Thus, in contrast to nuzzling behavior, male sex behavior appeared to be eliminated during androgen deprivation.     

Cell death and sexual differentiation of behavior: worms, flies, and mammals

Sex differences in the nervous system are found throughout the animal kingdom. Here, we discuss three prominent genetic models: nematodes, fruit flies, and mice. In all three, differential cell death is central to sexual differentiation and shared molecular mechanisms have been identified. Our knowledge of the precise function of neural sex differences lags behind. One fruitful approach to the 'function' question is to contrast sexual differentiation in standard laboratory animals with differentiation in species exhibiting unique social and reproductive organizations. Advanced genetic strategies are also addressing this question in worms and flies, and may soon be applicable to vertebrates.     

Comparison of Amplitude Recovery Dynamics of Two Limit Cycle Oscillator Models of the Human Circadian Pacemaker

At an organism level, the mammalian circadian pacemaker is a two‐dimensional system. For these two dimensions, phase (relative timing) and amplitude of the circadian pacemaker are commonly used. Both the phase and the amplitude (A) of the human circadian pacemaker can be observed within multiple physiological measures—including plasma cortisol, plasma melatonin, and core body temperature (CBT)—all of which are also used as markers of the circadian system. Although most previous work has concentrated on changes in phase of the circadian system, critically timed light exposure can significantly reduce the amplitude of the pacemaker. The rate at which the amplitude recovers to its equilibrium level after reduction can have physiological significance. Two mathematical models that describe the phase and amplitude dynamics of the pacemaker have been reported. These models are essentially equivalent in predictions of phase and in predictions of amplitude recovery for small changes from an equilibrium value (A=1), but are markedly different in the prediction of recovery rates when A<0.6. To determine which dynamic model best describes the amplitude recovery observed in experimental data; both models were fit to CBT data using a maximum likelihood procedure and compared using Akaike's Information Criterion (AIC). For all subjects, the model with the lower recovery rate provided a better fit to data in terms of AIC, supporting evidence that the amplitude recovery of the endogenous pacemaker is slow at low amplitudes. Experiments derived from model predictions are proposed to test the influence of low amplitude recovery on the physiological and neurobehavioral functions.     

Optimal stimulus shapes for neuronal excitation

An important problem in neuronal computation is to discern how features of stimuli control the timing of action potentials. One aspect of this problem is to determine how an action potential, or spike, can be elicited with the least energy cost, e.g., a minimal amount of applied current. Here we show in the Hodgkin & Huxley model of the action potential and in experiments on squid giant axons that: 1) spike generation in a neuron can be highly discriminatory for stimulus shape and 2) the optimal stimulus shape is dependent upon inputs to the neuron. We show how polarity and time course of post-synaptic currents determine which of these optimal stimulus shapes best excites the neuron. These results are obtained mathematically using the calculus of variations and experimentally using a stochastic search methodology. Our findings reveal a surprising complexity of computation at the single cell level that may be relevant for understanding optimization of signaling in neurons and neuronal networks.     

Regulation of motoneuron death in the spinal nucleus of the bulbocavernsus

A sexual dimorphism in the number of motoneurons in the spinal nucleus of the bulbocavernosus (SNB) of rats is engendered by a sex difference in ontogenetic cell death. Testicular secretions, specifically androgenic steroids, reduce SNB motoneuron death in males. The fate of the target muscles generally mirrors that of the motoneurons, and androgens appear to exert their effects upon the target muscles, sparing the motoneurons as a secondary consequence. Treatment with ciliary neurotrophic factor can also spare SNB motoneurons in newborn females, raising the possibility that this factor normally mediates androgen's effect upon motoneuron survival. The ontogeny of calcitonin gene-related peptide immunoreactivity is delayed in SNB cells compared with other motoneurons and is further delayed in the SNB cells of females. In both sexes, calcitonin gene-related peptide is detected after the period of SNB motoneuron death is complete. A sex difference in motoneuron number is also seen in the human homologue of the SNB and, because ontogenetic death of motoneurons in humans overlaps the period of androgen secretion, may arise in a manner similar to that in the rat SNB. © 1992 John Wiley & Sons, Inc.     

The spinal nucleus of the bulbocavernosus: firsts in androgen-dependent neural sex differences

Cell number in the spinal nucleus of the bulbocavernosus (SNB) of rats was the first neural sex difference shown to differentiate under the control of androgens, acting via classical intracellular androgen receptors. SNB motoneurons reside in the lumbar spinal cord and innervate striated muscles involved in copulation, including the bulbocavernosus (BC) and levator ani (LA). SNB cells are much larger and more numerous in males than in females, and the BC/LA target muscles are reduced or absent in females. The relative simplicity of this neuromuscular system has allowed for considerable progress in pinpointing sites of hormone action, and identifying the cellular bases for androgenic effects. It is now clear that androgens act at virtually every level of the SNB system, in development and throughout adult life. In this review we focus on effects of androgens on developmental cell death of SNB motoneurons and BC/LA muscles; the establishment and maintenance of SNB motoneuron soma size and dendritic length; BC/LA muscle morphology and physiology; and behaviors controlled by the SNB system. We also describe new data on neurotherapeutic effects of androgens on SNB motoneurons after injury in adulthood.