Contributions of placodal and neural crest cells to avian cranial peripheral ganglia

The method of embryonic tissue transplantation was used to confirm the dual origin of avian cranial sensory ganglia, to map precise locations of the anlagen of these sensory neurons, and to identify placodal and neural crest-derived neurons within ganglia. Segments of neural crest or strips of presumptive placodal ectoderm were excised from chick embryos and replaced with homologous tissues from quail embryos, whose cells contain a heterochromatin marker. Placode-derived neurons associated with cranial nerves V, VII, IX, and X are located distal to crest-derived neurons. The generally larger, embryonic placodal neurons are found in the distal portions of both lobes of the trigeminal ganglion, and in the geniculate, petrosal and nodose ganglia. Crest-derived neurons are found in the proximal trigeminal ganglion and in the combined proximal ganglion of cranial nerves IX and X. Neurons in the vestibular and acoustic ganglia of cranial nerve VIII derive from placodal ectoderm with the exception of a few neural crest-derived neurons localized to regions within the vestibular ganglion. Schwann sheath cells and satellite cells associated with all these ganglia originate from neural crest. The ganglionic anlagen are arranged in cranial to caudal sequence from the level of the mesencephalon through the third somite. Presumptive placodal ectoderm for the VIIIth, the Vth, and the VIIth, IXth, and Xth ganglia are located in a medial to lateral fashion during early stages of development reflecting, respectively, the dorsolateral, intermediate, and epibranchial positions of these neurogenic placodes.     

Quantitative distribution of NADPH-diaphorase-positive myenteric neurons in different segments of the developing chicken small intestine and colon

NADPH-diaphorase (NADPH-d) was used as a marker for neuronal nitric oxide synthase in order to investigate the nitrergic neurons of the developing myenteric ganglia on whole-mount preparations in the proximal and distal segments of the small intestine and in the colon of the chicken embryo, between incubation days 12 and 19. Neurons that were positive for NADPH-d were counted in randomly selected myenteric ganglia. The data obtained from each area and each age group were subjected to two-way analysis of variance (ANOVA) and the Student–Newman–Keuls test. Between incubation days 12 and 19, the originally narrow-meshed myenteric plexus with its high ganglionic density progressively became wide-meshed and the ganglionic density decreased significantly. Quantitative analysis further revealed a significant decrease in the NADPH-d-positive nerve cell density with age. At the same time, the constant or even increasing number of nitrergic cells per ganglion may indicate that the decreasing cell density may be a result of the growth of the bowel with decreasing ganglion density rather than a decrease in the total number of myenteric nitrergic cells. Regional differences in the dynamics of the quantitative changes were revealed. A significant decrease in the nitrergic cell number appeared earlier in the proximal than in the distal segments of the small intestine or in the colon. In contrast, the significant decline of the ganglionic density was first noticed in the colon at the same time.     

Reorganization of peptidergic systems during brain regeneration in Eisenia fetida (Oligochaeta, Annelida)

After the extirpation of the brain reorganization of the peptidergic (FMRFamide, neuropeptide Y, proctolin) systems was studied in the newly forming cerebral ganglion of the annelid Eisenia fetida. During regeneration, all immunoreactive fibres appear on the 1st-2nd postoperative day. At the beginning of regeneration, immunoreactive neurons and fibres form a mixed structure in the wound tissue. On the 3rd postoperative day, FMRFamide positive and neuropeptide Y-immunoreactive, while on the 7th postoperative day proctolin-immunoreactive neurons appear in the loose wound tissue. From the 25th postoperative day a capsule gradually develops around it. The neurons of the preganglion move to the surface of the newly appearing preganglion. The number of these cells gradually increase, and by the 72th-80th postoperative days the localization and number of peptide-immunoreactive neurons is similar to that in the intact one. The neurons of all examined peptidergic systems may originate from the neuroblasts, situated on the inner and outer surface of the intact ganglia (e.g. suboesophageal and ventral cord ganglia). In addition FMRFamide and proctolin immunoreactive neurons may take their derive by mitotic proliferation from the pharyngeal neurons, too.     

Development of satellite-neuron relations in the spinal ganglia of the rat

Cerebrospinal ganglia of 11-, 13-, 15-, 18- and 20-day-old embryos have been studied light- and electron-microscopically. On the 11th day of anlage the ganglia are presented exclusively as neuroblasts: the satellite-cells are formed later from lemmoblasts that install into the ganglion on the 13th-15th days both from the surrounding mesenchyme and from the developing roots and trunklets of the cerebrospinal nerves. In the ganglion they undergo proliferation and further differentiation. On the 15th day simple satellite-neuron interrelationships are formed: one satellite-cell is tightly connected with perikaryons of some neuroblasts and young neurons. On the 18th day around some neurons a double layered capsule is formed from the satellite-cells. Multilayered tunics are formed only on the 10th-15th day after birth. During the process of the capsule formation an unusual reactive character of the satellite-neuron interrelationships is revealed, it is manifested as, performed by the satellite-cells, an active microphagocytosis of the perikaryon cytoplasmic processes and the initial segment of the sensitive neuron main process. In embryogenesis the lemmocyte-axon interrelationships develop earlier than the satellite-neuronal ones.     

Embryonic origin of substance P containing neurons in cranial and spinal sensory ganglia of the avian embryo

The ontogeny of the neurons exhibiting substance P-like immunoreactivity (SPLI) was examined in the spinal and cranial sensory ganglia of chick and quail embryos. It was shown that in dorsal root ganglia (DRG) virtually all neuronal somas occupying the mediodorsal (MD) region of the ganglia are SPLI-positive while the larger neurons of the lateroventral (LV) area are SPLI-negative. In the cranial nerve ganglia, both types of neurons coexist in the trigeminal ganglion but with a different distribution: small neurons with SPLI are proximal while large neurons without SPLI occupy the maxillomandibular and ophthalmic lobes. The distal ganglia of nerves VII and IX (i.e., geniculate, petrosal) do not show cell bodies with SPLI in the two species considered. A few of them only (about 12%) are found in the nodose (distal ganglion of nerve X). The proximal ganglia of nerves IX and X (i.e., superior-jugular complex) are composed of small neurons which virtually all exhibit SPLI. Chimaeric cranial sensory ganglia were constructed by grafting the quail hind-brain primordium into chick embryos. Revelation of SPLI was combined with acridine orange staining on the same sections in order to ascertain the placodal (chick host) or neural crest (quail donor) origin of the SP-positive neurons in each type of ganglion. We found that all the neurons showing SPLI are derived from the neural crest in the trigeminal and in the superior and jugular ganglia. In the geniculate, petrosal, and nodose all the neurons are derived from the placodal ectoderm. The small number of SPLI-positive cells of the nodose ganglia are not an exception to this rule. Therefore, generally speaking, the sensory neurons of the cranial ganglia that express the SP phenotype are derived from the crest, with the exception of some neurons present in the nodose of both quail and chick embryos and which are of placodal origin. The vast majority of placode-derived neurons do not have amounts of SP that can be detected under the conditions of the present study.     

Regeneration of neuron processes of the spinal ganglion after its experimental section in rats

Sensory fibers regeneration after dorsal root ganglion (DRG) section was studied in the rat. After middle cross-section of DRG-13 (left side) there were neurons in the proximal part with damage to peripheral processes and in the distal part with damage to central processes. Axonal ionophoresis of cobalt salts was used for the study of sensory fibers regeneration through the scar in DRG during 3, 7, 15, 30, 120 and 180 days after the damage. Peculiarity regeneration of the sensory fibers was shown in spite of damage localization near ganglion cells body, the regeneration of peripheral and central processes of ganglion cells started already on the 3rd day, and sprouting sensory fibers through the scar of DRG--on the 7th day.     

Placodal sensory neurons in culture: nodose ganglion neurons are unresponsive to NGF, lack NGF receptors but are supported by a liver-derived neurotrophic factor

Explant and dissociated neuron-enriched cultures of nodose ganglia (inferior or distal sensory ganglion of the Xth cranial nerve) were established from chick embryos taken between embryonic Day 4 (E4) and Day 16 (E16). The response of each type of culture to nerve growth factor (NGF) was examined over this developmental range. At the earliest ages taken (E4-E6), NGF elicited modest neurite outgrowth from ganglion explants cultured in collagen gel for 24 hr, although the effect of NGF on ganglia taken from E4 chicks was only marginally greater than spontaneous neurite extension from control ganglia of the same developmental age. The response of nodose explants to NGF was maximal at E6-E7, but declined to a negligible level in ganglia taken from E9-E10 or older chick embryos. In dissociated neuron-enriched cultures, nodose ganglion neurons were unresponsive to NGF throughtout the entire developmental age range between E5 and E12. In contrast to the lack of effect of NGF, up to 50% of nodose ganglion neurons survived and produced extensive neurites in dissociated cultures, on either collagen- or polylysine-coated substrates, in the presence of extracts of late embryonic or early posthatched chick liver (E18-P7). Antiserum to mouse NGF did not block the neurotrophic activity of chick (or rat or bovine) liver extracts. Whether cultured with chick liver extract alone or with chick liver extract plus NGF, nodose ganglion neurons taken from E6-E12 chick embryos and maintained in culture for 2 days were devoid of NGF receptors, as assessed by autoradiography of cultures incubated with 125I-NGF. Under similar conditions 70-95% of spinal sensory neurons (dorsal root ganglion--DRG) were heavily labeled. 2+     

Development of substance P and neurokinin A immunoreactivity in ganglia supplying nerves to the submandibular glands of the rat

Developing submandibular, trigeminal and superior cervical ganglia, which provide innervation to the submandibular glands, were studied for substance P (SP)-and neurokinin A (NKA)-immunoreactive (IR) ganglion cells and nerve fibres in rat. These ganglia were examined by using an indirect immunofluorescence technique at daily intervals from the 16th day in utero (i.u.) until birth, and subsequently on the 2nd, 5th, 7th, 12th, 16th, 30th, 42nd postnatal day and in the adult (3 months). In the submandibular ganglion SP- and NKA-IR cells and fibres first appeared in considerable numbers on the 19th day i.u. (in one sample out of five on the 18th day i.u.), when more than 90% of the ganglion cells were immunoreactive to SP and NKA. The number stayed at more than 90% to the 7th postnatal day and then slowly decreased to the levels of adult animals (18% SP, 17% NKA). The first SP- and NKA-IR ganglion cells and fibres appeared in the trigeminal ganglion on the 18th day i.u. when they represented 7% (SP) and 4% (NKA) of the ganglion cells. The number of SP- and NKA-IR cells increased steadily, reaching a maximum at the time of birth when 68% (SP) and 74% (NKA) of the ganglion cells were immunoreactive. Thereafter they began to decrease toward the level of an adult rat (10% SP, 11% NKA). In the superior cervical ganglion only a few SP-and NKA-IR ganglion cells were detected from the 19th day i.u. to the fifth postnatal day. Positive ganglion cells were also occasionally found in the nerve trunks outside the superior cervical ganglion. From the seventh day onwards no SP- or NKA-IR ganglion cells were found. SP-and NKA-IR SIF (small intensively fluorescent) cells were detected from the 16th postnatal day onwards.     

DEVELOPMENTAL CHANGES OF CHOLINESTERASES AND MONOAMINE OXIDASE IN CHICK EMBRYO SPINAL AND SYMPATHETIC GANGLIA

Abstract— Total cholinesterase, acetylcholinesterase, (AChE) and monoamine oxidase (MAO) activity and protein content were determined throughout the embryonic life of the chick in spinal and sympathetic ganglia. The greatest part of total cholinesterase activity was due to AChE.
AChE and MAO activity increased in both spinal and sympathetic ganglia very similarly from the 6th to the 12th day of incubation; from this day on a significant divergence occurred, mainly owing to a steady fall in spinal ganglion AChE, which decreased to approximately one tenth of the maximum value. The ratio of MAO activity in sympathetic and spinal ganglia increased from the 8th day onwards and approached 5·0 at hatching. The ratio between sympathetic and spinal ganglia, for AChE, choline acetylase (ChAc) and MAO activity, suggests a relationship between the maturation of the synapse in the sympathetic ganglia and the maximal activity of these enzymes.     

Effect of ethanol on the cerebellar cortex of the chick embryo

The effect of ethanol on the cerebellar cortex of chick embryos was studied in semi-thin sections of material prepared for electron microscopy. The embryos were injected with ethanol on the 3rd or 6th day of incubation and observed until days 13, 15, 17 and 21 of development. A decrease was seen in the number of germinal cells generated, together with defects in neuronal migration and the existence of a lower quantity of cells due to a generalised process of cell death. At the same time, a progressive neuronal degeneration was observed until the 15th day of incubation, the tissue recovering progressively on days 17 and 21. On the other hand, the embryos treated with ethanol on the 3rd day were less affected than those injected on the 6th day.     

Reorganization of monoaminergic systems in the earthworm,Eisenia fetida,following brain extirpation

The present study describes the major aspects of how monoaminergic (serotonin, dopamine) systems change in the course of regeneration of the brain in the earthworm (Eisenia fetida), investigated by immunocytochemistry, HPLC assay, and ligand binding. Following brain extirpation, the total regeneration time is about 80 days at 10 degrees C. On the 3rd postoperative day serotonin, and on the 11th postoperative day tyrosine hydroxylase-immunoreactive neurons can be observed in the wound tissue. Thereafter the number of the immunoreactive cells increases gradually, and by the 76th-80th postoperative days all serotonin- and tyrosine hydroxylase-immunopositive neurons can be found in their final positions, similarly to those observed in the intact brain. Labeled neurons located in the dorsal part of the regenerated brain appear earlier than the cells in lateral and ventral positions. Both serotonin- and tyrosine hydroxylase-immunoreactive neurons of the newly formed brain seem to originate from undifferentiated neuroblasts situated within and around the ventral ganglia and the pleura. Dopaminergic (tyrosine hydroxylase-immunoreactive) elements may additionally derive from the proliferation of neurons localized in the subesophageal ganglion and the pharyngeal nerve plexus. Following brain extirpation, both serotonin and dopamine levels, assayed by HPLC, first increase in the subesophageal ganglion; by the 25th day of regeneration, the monoamine content decreases in it and increases in the brain. Hence it is suggested that monoamines are at least partly transported from this ganglion to the regenerating brain. At the same time, (3)H-LSD binding can be detected in the regenerating brain from the 3rd postoperative day, showing a continuous increase until the 80th postoperative day, suggesting a guiding role of postsynaptic elements in the monoaminergic reinnervation of the newly formed brain.     

Preparation of the spinal neurons and their bushy receptors for morphophysiological study

A preparation of the sensory neuron of the spinal ganglion with dendritic processes for simultaneous morphological and physiological investigations is described. It consists of a frog urinary bladder with bushy interoceptors in its wall, two vesicle neurons, two abdominal branches of the X spinal nerves and two IX spinal ganglia with ventral and dorsal roots branching off from them. The total length from the receptors to the ganglion neurons is 20-30 mm. In the ganglia a zone of the neuronal bodies localization is found, their processes form receptors; the zone includes as many as 9 neurons, 50-80 mkm in size. A vital fine structure of the ganglion cells and their satellites is traced. There are three types of cells in the ganglion--large, middle and small. Electrophysiological control has demonstrated that the preparation is viable for several hours.     

Developmental potentialities in the nonneuronal population of quail sensory ganglia

Sensory ganglia taken from quail embryos at E4 to E7 were back-transplanted into the vagal neural crest migration pathway (i.e., at the level of somites 1 to 6) of 8- to 10-somite stage chick embryos. Three types of sensory ganglia were used: (i) proximal ganglia of cranial sensory nerves IX and X forming the jugular-superior ganglionic complex, whose neurons and nonneuronal cells both arise from the neural crest; (ii) distal ganglia of the same nerves, i.e., the petrosal and nodose ganglia in which the neurons originate from epibranchial placodes and the nonneuronal cells from the neural crest; (iii) dorsal root ganglia taken in the truncal region between the fore- and hindlimb levels. The question raised was whether cells from the graft would be able to yield the neural crest derivatives normally arising from the hindbrain and vagal crest, such as carotid body type I and II cells, enteric ganglia, Schwann cells located along the local nerves, and the nonneuronal contingent of cells in the host nodose ganglion. All the grafted cephalic ganglia provided the host with the complete array of these cell types. In contrast, grafted dorsal root ganglion cells gave rise only to carotid body type I and II cells, to the nonneuronal cells of the nodose ganglion, and to Schwann cells; the ganglion-derived cells did not invade the gut and therefore failed to contribute to the host's enteric neuronal system. Coculture on the chorioallantoic membrane of aneural chick gut directly associated with quail sensory ganglia essentially reinforced these results. These data demonstrate that the capacity of peripheral ganglia to provide enteric plexuses varies according to the level of the neuraxis from which they originate.     

Pharmacological characteristics of three silent giant neurons, v-LPSN, v-1-VOrN and v-VLN, identified on the ventral surface in the left parietal and the visceral ganglia of the suboesophageal ganglia of an African giant snail (Achatina fulica Férussac)

Three giant neurons, named v-LPSN, v-1-VOrN and v-VLN, were identified on the ventral surface of the left parietal ganglion and the visceral ganglion in the suboesophageal ganglia of an African giant snail (Achatina fulica Férussac). They lacked the spontaneous spike discharges in the normal state. The pharmacological features of the three neurons, in relation to the principal common neurotransmitter substances and their derivatives were examined. The v-LPSN (ventral-left parietal silent neuron) (diameter: about 130 microns) is situated in the left parietal ganglion close to the visceral ganglion. The neuron was excited by histamine (the minimum effective concentration [MEC]: 3 X 10(-4) M) and inhibited slightly by acetylcholine (Ach) and its related substances. The v-1-VOrN (ventral-left-visceral oral neuron) (diameter: about 130 microns) is situated in the left and oral part of the visceral ganglion. The neuron was inhibited markedly by dopamine (DA) (MEC: 3 X 10(-4) M) and slightly by Ach and its related substances. No substance producing a marked excitation of the neuron has been found yet. The v-VLN (ventral-visceral large neuron) (diameter: about 300 microns) is found in the centre of the visceral ganglion. The neuron was excited by L-norepinephrine (MEC: 10(-4) M), DL-octopamine (MEC: 2 X 10(-4) M), 5-hydroxytryptamine (MEC: 10(-4) M) and beta-hydroxy-L-glutamic acid (MEC: 10(-4) M) and inhibited by DA (MEC: 10(-4) M), GABA (MEC: 3 X 10(-5) M) and Ach (MEC: 10(-4) M). L-Epinephrine showed varied effects (MEC: 10(-4) M), which were either excitatory or inhibitory.     

The level of nerve growth factor (NGF) as a function of innervation : A correlative radio-immunoassay and bioassay study of the rat iris

A two-site radio-immunoassay for βNGF demonstrated 5–10 pg of NGF in the normal, adult rat iris. Ciliarectomy or sympathectomy did not significantly alter the amount of NGF after 10 days. However, denervation including all sensory axons (stereotactic lesion distal to the trigeminal ganglion) increased the level to about 100 pg of NGF. Total denervation resulting from homologous transplantation of the iris gave a similar increase after only 2 days. Fibre outgrowth responses evoked by corresponding iris explants in an NGF bioassay supported the results and suggested in addition that sympathetic denervation may cause a moderate transient increase in NGF after 3 days. It seems that sensory nerves in particular influence the level of NGF in a terminal field, either by a high capacity for uptake and removal of NGF or by exerting a negative feed-back on the production or processing of this growth factor.     

Various aspects of organogenesis of the colon in Gallus domesticus: morphologic observation

The authors have investigated the morphological aspects of the wall components of the developing colon in the chick embryo (Gallus domesticus) from the 7th to th 15th day of incubation. Particular attention has been given to the lumen recanalization, phenomenon which occurs also in other animal species. The most significant results can be summarized as follows: 1) the lumen is recanalized at the 7th day only at the proximal part of the colon (Fig. 1, Tav. 1), while at the distal tract it is still completely filled by an epithelial plug (Fig. 2, Tav. 1). Therefore the recanalization of the lumen takes place cranio-caudad. 2) At the 8th day the process of recanalization of the lumen shows, in the distal part of the colon, well defined modalities. Radially oriented intraepithelial spaces within the epithelium filling the lumen join other semilunar intercellular spaces, which are placed near the central part of the occluded lumen (Fig. 3). By the junction of a couple of radially oriented spaces with one semilunar space, an U-shaped intercellular space derives, which delimits an incoming epithelial fold (Tav. 3). Such a phenomenon is continued also during the 9th and 10th day of incubation (Fig. 6, Tav. 2). 3) At the 11th day the colonic lumen is completely open and, in its distal part, the appearance of the primordial previllous ridges can be observed (Fig. 7). In the proximal tract the previllous ridges develop one day later (Fig. 8). 4) At the 13th day, in the distal part of the colon, the first appearance of crypts occurs (Fig. 10). So, while the process of recanalization of the lumen is cranio-caudad, the formation of previllous ridges and crypts proceeds caudo-cranially. 5) From the 11th day onwards the lamina propria is actively involved in the process of formation of the previllous ridges. Only at the 14th day, in the distal part of the colon anlage, the appearance of the muscularis mucosae is observed (Fig. 11). 6) The muscle layer and the subserous stratum do not show appreciable morphological changes in the course the considered period of incubation.     

Quantitative in vitro studies on the nerve growth factor (NGF) requirement of neurons. II. Sensory neurons

Studies were carried out in dissociated cell cultures on the nerve growth factor (NGF) requirement of chick embryo dorsal root ganglionic (DRG) neurons. Findings were: (i) The minimum level of 2.5 S NGF required to sustain the survival of maximal numbers of process-bearing cells derived from 8-day (E8) embryonic DRGs is 0.5 ng/ml (～2 × 10^?11M). (ii) Cultures derived from chick embryos of increasing ages (E8 to E18) showed a progressive increase in the proportion of process-bearing cells which survived in the absence of NGF. While few process-bearing cells survived in cultures of E8 ganglia in the absence of NGF, survival of neurons in cultures derived from E17 and E18 ganglia was not affected by the absence of the factor. Comparable results were obtained with cultures in which the number of non-neuronal cells was greatly reduced. (iii) Neurons derived from E8 ganglia lost their NGF requirement in culture at a conceptual age similar to that which they appear to do so in vivo. These results are discussed with respect to the role of NGF in development of sensory neurons.     

Postembryonic development of serotonin-immunoreactive neurons in the central nervous system of the blowfly

Summary Neurons immunoreactive with antibodies to serotonin (5-HT) were mapped in the thoracico-abdominal ganglia of the blowfly, Calliphora erythrocephala, during postembryonic development. Reconstructions from serial sections of tissue processed with a preincubation PAP-method permitted a detailed analysis of the morphological changes occurring in 5-HT-immunoreactive (5-HTi) neurons.All the 5-HTi cell bodies in the thoracico-abdominal ganglia of the 3rd instar larva, except two in the metathoracic ganglion, retain their immunochemical phenotype throughout pupal development. Hence, all the adult 5-HTi neurons in these ganglia differentiate during embryonic development. The finer processes of the larval 5-HTi neurons undergo a substantial regression during the first 24 h of pupal development, and thereafter new branches form on the primary processes of the same cell bodies. The slight change in relative position of 5-HTi cell bodies and the reorganization of the neuropil into an adult pattern occur during the first half of pupal development. The neuropil mass and extent of 5-HTi processes continue to increase during the following days and appear to be fully developed two days (80% of pupal development) before hatching.On the basis of the presented data, some of the basic processes are discussed that lead to the transformation of the larval nervous system into its adult form.     

The Ability of Diphenylpiperazines to Prevent Neuronal Death in Dorsal Root Ganglion Neurons In Vitro After Nerve Growth Factor Deprivation and In Vivo After Axotomy

Abstract: The mechanism of neuroprotection by the calcium channel antagonist flunarizine against neuronal death is unknown. We investigated the ability of other calcium channel antagonists (cinnarizine, nimodipine, nicardipine, diltiazem, and verapamil), calmodulin antagonists, and calpain inhibitors to prevent neuronal death in rat dorsal root ganglion neurons in vitro after nerve growth factor (NGF) deprivation and the ability of cinnarizine and diltiazem to protect in vivo after axotomy. In vitro, only neurons treated with cinnarizine or flunarizine were protected from death after withdrawal. In vivo, cinnarizine, but not diltiazem, protected dorsal root ganglion neurons in rats after unilateral sciatic nerve crush. Intracellular calcium concentration ([Ca²⁺],) was evaluated with fura 2 after NGF deprivation In vitro. Neurons "committed to die" 24 h after NGF deprivation displayed a decline in [Ca^a+], before visible morphological deterioration consistent with cell death. The influx of extracellular calcium was not necessary to produce neuronal death. Neurons deprived of NGF gradually lost the ability to respond to elevated external potassium with an increase in [Ca²⁺], during the first 24 h after trophic factor deprivation. After 24 h, neurons deprived of NGF could not be rescued by readministration of NGF. Neurons protected from cell death with diphenylpiperazines maintained their response to high external potassium, suggesting continued membrane integrity. We speculate that diphenylpiperazines may protect sensory neurons via an unknown mechanism that stabilizes cell membranes.     

Trophic effects of androgen: Development and hormonal regulation of neuron number in a sexually dimorphic vocal motor nucleus

In Xenopus laevis, the laryngeal motor nucleus (n. of cranial nerves IX‐X) is part of a sexually differentiated, androgen sensitive neuromuscular system devoted to vocalization. Adult males have more n. IX‐X neurons than females; however, during development of n. IX‐X, the rate of neurogenesis does not appear to differ between the sexes. In this study, we explored the role of naturally occurring cell death in the development of this nucleus and asked whether cell death might be involved in establishing the sex difference in neuron number. Counts of n. IX‐X neurons reveal that at tadpole stage 56, males and females have similar numbers of n. IX‐X neurons, but by stage 64 male neuron numbers are greater. This sex difference arises owing to a greater net loss of neurons in females—males lose ∼25% of their n. IX‐X neurons between stages 56 and 64, while females lose ∼47%. Sexual differentiation of n. IX‐X neuron number coincides with a period of developmental cell death, as evidenced by terminal transferase‐mediated dUTP nick‐end labeling and the presence of pyknotic nuclei in n. IX‐X. A role for gonadal hormones in controlling cell number was examined by treating tadpoles with exogenous androgen and determining the number of n. IX‐X neurons at stage 64. Dihydrotestosterone (DHT) treatment from the beginning of the cell death period (stage 54) until stage 64 had no effect on the number of n. IX‐X neurons in males but did significantly increase n. IX‐X neuron number in females. This increase was sufficient to abolish the sex difference normally observed at stage 64. Although DHT induced increases in female neuron number, it did not induce increases in cell proliferation or addition of newly born neurons to n. IX‐X. DHT may therefore have increased neuron number by protecting cells from death. We conclude that androgens can influence the survival of n. IX‐X neurons during a period of naturally occurring cell death, and that this action of androgen is critical to the development of sex differences in n. IX‐X neuron number. © 1999 John Wiley & Sons, Inc. J Neurobiol 40: 375–385, 1999