On variables that affect estimates of the true sizes and densities of radioactively labeled cell nuclei

Clark, Stephen J.; Cynx, Jeffrey; Alvarez-Buylla, Arturo; O'loughlin, B.; Nottebohm, Fernando

doi:10.1002/cne.903010111

Cited by 20 publications

(11 citation statements)

References 12 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Counts of 3H-labeled cells with nuclear diameters within the range we encountered, using tissue sections 6 ,um thick, are very unlikely to result in over or under estimates of the true number of these cells. In fact, all of our 3H-labeled neurons, whether their mean diameter was close to 7 or 9 um, were probably equally likely to be counted (50). In accordance with the above, there was no simple relation between the nuclear size and the number of 3H-labeled neurons in the different experimental groups.…”

Section: Discussionsupporting

confidence: 63%

The life span of new neurons in a song control nucleus of the adult canary brain depends on time of year when these cells are born.

Nottebohm

O'loughlin²,

Gould³

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

The number of high vocal center (HVC) neurons labeled in adult male caries by systemic Injections of [3H]thymidine depended on season and survival time. This was true for HVC neurons projecting to the robust nucleus of the archistriatum and for other HVC neurons that could not be retrogradely filled from the robust nucleus of the archistriatum. Birds Injected in October and killed 40 days later had twice as many labeled HVC neurons as birds injected in May and killed 40 days later. However, this difference became much larger (5 times) when the birds were allowed to survive for 4 months. Whereas more than half of the spring-born neurons disappeared between 40 days and 4 months, there was no reduction in the number of fall-born neurons present at the 4-month survival point. We infer that seasonal variables affect the life span of HVC neurons born in adulthood.It has been known for many years that the developing nervous system overproduces neurons. Many parts of the developing central nervous system retain only a fraction of the neurons originally produced (1-5). The conditions that determine this selective death are under intensive study (e.g., refs. 6-9). Neuronal death is also seen in some diseases ofthe adult human brain-e.g., Alzheimer and Parkinson diseases. Several studies (e.g., refs. 10-12) have looked at the factors or conditions that may prevent or delay neuronal death during development or in the adult diseased brain. There is hope that factors that prevent selective death during development may also rescue neurons that die during disease. But such studies need not be restricted to the developing brain. Neurogenesis and neuronal replacement occur spontaneously in a part of the song system of adult songbirds. This offers the opportunity to study neuronal death and replacement in a healthy adult vertebrate brain and in a system that controls a well-characterized behavior.Oscine songbirds acquire their song by reference to auditory information (13)(14)(15). The circuits involved in the acquisition and production of learned song have been described (16-19). One ofthe telencephalic nuclei, the high vocal center (HVC), necessary for the production of learned song (16,20,21), is of particular interest because it incorporates and replaces neurons in adulthood (22)(23)(24)(25)(26). More than half of the adult-formed neurons added to the HVC (26-28) grow an axon that reaches nucleus robustus archistriatalis (RA), on the efferent pathway for song production (16). The number of HVC neurons, including those that project to RA, ceases to increase in adulthood and neurogenesis in this system is part of a process of neuronal replacement (25,28,29).Earlier work (27) GBq; New England Nuclear). These birds received the PHIthymidine treatment at two times of the year, May (spring) or September (fall), and were divided into four groups as described in Table 1. All birds were weighed, anesthetized with 50 A1 of a dilution of Nembutal (Abbott; 10 mg/ml), and given FluoroGold [2% (wt/vol) solution in saline] microinj...

show abstract

Section: Discussionsupporting

confidence: 63%

The life span of new neurons in a song control nucleus of the adult canary brain depends on time of year when these cells are born.

Nottebohm

O'loughlin²,

Gould³

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…We counted as new neurons cells that, in addition to their neuronal staining, had a number of exposed silver grains over their nucleus that was at least ϫ20 above background. The very stringent criterion for positive identification of a labeled neuron we used tended to ignore nuclear diameters of Ͻ5 m. For this reason, and taking into account the range of diameters observed and thickness of sections, we did not correct new neuron counts for cell splitting (16). Had these corrections been used, they would have been small and would not have changed the results reported here.…”

Section: Methodsmentioning

confidence: 99%

Timing of brain-derived neurotrophic factor exposure affects life expectancy of new neurons

Alvarez-Borda

Haripal

Nottebohm

2004

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

The high vocal center (HVC) of adult male canaries, Serinus canaria, is necessary for the production of learned song. New neurons are added to HVC every day, where they replace older neurons that have died, but the length of their survival depends on the time of year when they are born. A great number of HVC neurons born in the fall, when adult canaries learn a new song, are still present 8 mo later, when this song is used during the breeding season. By contrast, most of the neurons born in HVC in the spring, when little song learning takes place, disappear much sooner. Here we show that infusion of brain-derived neurotrophic factor into HVC during days 14 -20 after new HVC neurons are born in the spring confers on them a life expectancy comparable to that of fall-born neurons; this extension on life is not seen when infusion occurs 10 days earlier or later. We suggest that there is, in the adult HVC, a subset of neurons whose life expectancy is determined by brain-derived neurotrophic factor during a sensitive period soon after these neurons reach destination and start forming connections.N ew neurons are born in the adult canary brain in the ventricular zone lining the wall of the lateral ventricles (1, 2). They migrate into the high vocal center (HVC) and assume a sedentary phenotype 8-15 days after their birth. The number of these new neurons is much reduced during their third week of life (3). By day 30, the axons of many of the surviving new HVC neurons have reached their target, and many of the new cells have become a functional part of existing circuits (3-5). Thereafter, the life expectancy of the neurons depends on the time of year when they are born. Whereas the number of fall-born neurons is virtually the same 30 and 120 days after their birth, this number drops by one-half at the later survival time in spring-born neurons (6). However, the mechanism that regulates this season-dependent survival͞attrition has not been worked out in detail.In addition to season, the presence of new neurons in HVC has been shown to depend on variables such as amount of singing (7,8), auditory experience (9), blood testosterone levels (8, 10), and the presence of brain-derived neurotrophic factor (BDNF) in HVC (11). Some of these factors are related. For example, both amount of singing and blood testosterone levels change seasonally (12) and both influence the amount of BDNF present in HVC (8). There are at least two major sources of BDNF in HVC: neurons that project to the robust nucleus of the archistriatum (7) and endothelial cells lining HVC capillaries (13); in addition, a small percentage of HVC neurons that project to area X also express BDNF (7). Knowing that BDNF can influence neuron survival and that there is a dramatic decline in new neuron numbers 15-22 days after their birth (3), we decided to test whether BDNF infusion during this period could enhance the survival of new neurons born in the spring. We found that, whereas BDNF infusion into HVC during days 14-20 after new neurons are born markedly extends t...

show abstract

“…With 3H as the radioactive isotope, brain sections must be exposed to emulsion for weeks to months before adequate numbers of silver grains are reduced in the emulsion to allow discrimination of labeled cells. 0-Particles emitted by the decay of 3H penetrate poorly through more than 3 pm of tissue (Clark et al, 1990;Arnold, 1981 Quarmby et al, 1990). Application of ICC to study ARs in the avian song control system, however, has been hampered by the lack of an antibody that selectively labels ARs in songbird brains.…”

Section: Introductionmentioning

confidence: 99%

Use of PG-21 immunocytochemistry to detect androgen receptors in the songbird brain.

Smith

Brenowitz

Prins

1996

J Histochem Cytochem.

View full text Add to dashboard Cite

The avian song control system is an excellent model in which to study the effects of gonadal steroid hormones on neural and behavioral plasticity. Several of the brain regions that control song behavior concentrate androgens andlor estrogens. Investigations of the distribution and regulation of androgen receptors have been limited by the lack of a reliable immunocytochemical method to detect androgen receptors in the songbird brain. We describe a protocol by which the PG-21 polyclonal antibody to the rat androgen receptor can be used to label androgen receptor-containing cells in the songbird brain. By treating songbirds of several species with testosterone 90 min before sauifce and by using relatively IntroductionThe neural circuit that regulates song behavior in songbirds has emerged as a leading model for studying the effects of gonadal steroid hormones on brain structure and function. Song is a learned behavior that is sexually dimorphic in most species. The morphology of the brain nuclei that control song differs between sexes and between seasons (reviewed in Brenowitt and Kroodsma, in press;Arnold, 1992;DeVoogd, 1991;Konishi, 1989;Nottebohm, 1987).Both the organization and the activation of the song control system are strongly influenced by gonadal steroid hormones. Synergistic interactions of androgens and estrogens are essential for development of song control circuits in the brain and for production of song (reviewed in Brenowitz and K " a , in press). Song learning is influenced by gonadal steroids (Korsia and Bottjer, 1991;Marler et al., 1988). These hormones also play an important role in mediating seasonal plasticity of song nuclei in adult songbirds (Smith et al., 1993).Steroid autoradiography has been the primary tool used to study the distribution and regulation of gonadal steroid receptors in the song control system (Arnold et al., 1976;Zigmond et al., 1973). Autoradiographic studies of zebra finches (Zeniopygiaguttuta) and canaries (Serinur canariu) using [3H]-dihydrotestosterone (DHT) as a ligand indicate that androgen receptor-containing (AR+ ) cells are present in five song nuclei: the higher vocal center (HVC), the magnocellular nucleus of the anterior neostriatum (MAN), the robust nucleus of the archistriatum (RA), the intercollicular nucleus (ICo), and the tracheosyringeal portion of the hypoglossal nucleus (nXIIts) (Gahr, 1990;Nordeen et al., 1986). Androgen accumulation is also observed in the preoptic area, the magnocellular paraventricular nucleus, and the tuberal nucleus in the hypothalamus of these species '(Gahr, 1990; Nordeeti et al., 1986).The autoradiographic technique has several limitations as a means of studying the distribution of cells that contain AR. With 3H as the radioactive isotope, brain sections must be exposed to emulsion for weeks to months before adequate numbers of silver grains are reduced in the emulsion to allow discrimination of labeled cells. 0-Particles emitted by the decay of 3H penetrate poorly through more than 3 pm of tissue (Clark et al., 1990;Arnold, ...

show abstract

On variables that affect estimates of the true sizes and densities of radioactively labeled cell nuclei

Cited by 20 publications

References 12 publications

The life span of new neurons in a song control nucleus of the adult canary brain depends on time of year when these cells are born.

The life span of new neurons in a song control nucleus of the adult canary brain depends on time of year when these cells are born.

Timing of brain-derived neurotrophic factor exposure affects life expectancy of new neurons

Use of PG-21 immunocytochemistry to detect androgen receptors in the songbird brain.

Contact Info

Product

Resources

About