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PubMed2004

[Distribution of satellite DNA TkS1 in genomes of salamanders (Salamandridae)].

Genomes of 22 species and subspecies of salamandrids (Triturus, Cynops, Neurergus, Notophthalmus, Pachytriton, and Pleurodeles ) were studied. Satellite TkS1 (32-33 bp) was found in all examined species, both in direct and reversed orientations. The use of single-primer PCR with oligonucleotides, which are homologous to the most conservative part of satellite TkS1, allowed us to reveal DNA fragments, flanked by single copies (or small tandem repeats) of satellites TkS1 having various (5' and 3') orientation. Such fragments were observed in Pleurodeles waltl, Triturus a. alpestris, T. vulgaris lantzi, T. v. vulgaris, Neurergus crocatus and T. vulgaris graecus. The length of amplified DNA fragments in three subspecies of T. vulgaris differed. This might be connected with unequal amounts and different distribution of 5' and 3' copies of satellite TkS1 in their genomes. Patterns of DNA amplification in Triturus a. alpestris and Neurergus crocatus were quite similar. Only two species (Triturus a. alpestris and T. v. vulgaris) had approximately equal amounts and similar distribution of 5' and 3' copies of satellite TkS1 in their genomes.

Animals↗
PubMed1990

The spatial distribution of excitatory and inhibitory inputs to ganglion cell dendrites in the tiger salamander retina.

In response to focal stimuli, ganglion cell dendrites receive excitation over a relatively narrow extent of the inner plexiform layer (IPL). This excitation is embedded in 2 wider lateral inhibitory regions. Here we estimate the lateral dimensions of the inhibitory regions. Ganglion cells were whole-cell patch-clamped and dendrites were identified and located in retinal slices using Lucifer yellow in the pipettes. The spatial distribution of ganglion cell dendritic sensitivity was measured with puffs of transmitter substances applied at different distances along the dendrites. All ganglion cell dendrites were sensitive to glutamate, GABA, and glycine across their full extent. The responses to puffs decreased with lateral distance from the soma and were well fit by Gaussians. The responses to puffs of potassium showed a similar decrement with distance. Since potassium channels are probably uniformly distributed along the dendrites, the similarity in profiles suggests that receptor density is also uniform along the dendrites. The spatial distribution of responses of ganglion cells to excitatory and inhibitory synaptic inputs was measured by depolarizing local populations of bipolar terminals (and subsequently local populations of amacrine cells) with transretinal current (TRC). TRC-stimulating electrodes were displaced laterally, with respect to the ganglion cell soma, to generate response profiles. We estimated the dimensions of the inhibitory and excitatory signals received by the ganglion cells by removing the contributions of their dendrites, the stimulus, and other interneurons from the response profiles. The excitatory signal extended less than 100 microns, the approximate dimensions of the ganglion cell dendrites, and corresponds roughly to the width of the bipolar inputs. The GABAergic signal extended, on average, 253 microns and glycinergic signal extended, on average, 315 microns. These inhibitory signal dimensions correspond to the width of classes of amacrine cell processes measured in other studies.

Animals↗
PubMed1990

Differential germ cell proliferation in the salamander pleurodeles waltl: controls by sexual genotype and by thermal epigenetic factor before differentiation of sexual phenotype of gonads.

In the acceptance that, during early gonadogenesis, variations of germ cell (GC) proliferation express interactions between germ and somatic cells, early events occurring before histological differentiation of gonadal sex has been detected and timed through GC counts on larvae of Pleurodeles waltl (urodele amphibia) issued from male ZZ or female ZW monosexual offspring. Gonads differentiate in accordance with sexual genotype in ZZ and ZW larvae at room temperature and in ZZ larvae at 32 degrees C whereas they are sex-reversed at 32 degrees C in ZW larvae, becoming phenotypic neomales. At both the rearing temperatures, in genital ridges, GCs do not proliferate during a period called P0 period ending earlier in ZZ than in ZW larvae. The time when proliferation starts depends on sexual genotypes and determines a ZZP0 period shorter than ZWP0 period. After P0 period, at room temperature, a moderate increase in GC number determining a P1 period is observed in both ZZ and ZW larvae, whereas a strong proliferation, determining a P2 period, occurs on a differential pattern in ZZ and ZW larvae; thus, before sexual differentiation of gonads, ZW females have more GCs than ZZ males. At 32 degrees C, GC proliferation is moderate during P1 period and does not accelerate during P2 period in ZW larvae differentiating neotestes; they have a lower GC number than ZZ larvae reared at 32 degrees C. Thus, during P2 period, at both room temperature and at 32 degrees C, GC number correlates with future phenotype of gonads. Results suggest that differential molecular events arise during early gonadogenesis and that testes may differentiate in different ways according to whether phenotype conforms to genotype or sex reversion occurs.

Animals↗
PubMed1989

Primordial germ cell proliferation in the salamander Pleurodeles waltl: genetic control before gonadal differentiation.

PGC counts were carried out on larvae of Pleurodeles waltl (urodele amphibia) issued from standard, monosexual male and monosexual female offspring while the genital ridges were settling. During this period, which is characterized by a zero mitotic index (and is therefore called the Po period), and which lasts from stage 35 to stage 41, no PGC proliferation occurs. A statistical analysis indicated that PGC counts per larva are sex genotype independent and that offspring may be divided into three groups with average PGC counts of 96.9, 51.0 and 31.1, respectively. A fourth group with an average of 18.3 PGCs has been identified using experimental larvae reared at 30 degrees C from stage 30. The PGC count of 96.9 would result from at least three mitotic cycles. Before the Po period, germ cells are not identifiable. A hypothesis concerning genetic control of PGC proliferation before Po was deduced from this analysis.

Animals↗
PubMed1989

Amacrine cell interactions underlying the response to change in the tiger salamander retina.

The neural circuitry and pharmacology underlying transient signal formation at the bipolar-amacrine cell interface were studied. Synaptic currents were measured with whole cell patch clamp in retinal slices. Cell types were identified with Lucifer yellow staining. Activity was initiated with puffs of kainate of known time course and spatial spread delivered at bipolar dendrites. OFF bipolar cells responded to kainate with a sustained inward current, but ON bipolar cells were silent. Two types of amacrine cell were found: (1) narrow field cells, with processes that extended laterally less than 200 microns, responding with a sustained inward current, and (2) wide field cells, with processes that extended laterally by up to 1 mm, responding with a brief transient inward current followed by a more sustained outward current. We pharmacologically dissected the synaptic interactions underlying the transient current in the wide field amacrine cell. In the presence of 5-aminovaleric acid (AVA), the time course of this transient current was increased so that it resembled the response of bipolar cells. Because AVA is a GABAB antagonist, it appears to block an opposing signal that truncates the sustained excitatory bipolar input, thereby generating the transient. GABAB specificity is confirmed by (1) block of the transient inward current by baclofen, a GABAB agonist, and (2) block of the baclofen effect by AVA. The site of GABAB action appears to be presynaptic to the amacrine cell membrane because neither baclofen nor AVA, in combination with picrotoxin, had a direct effect at the amacrine cell membrane. GABAB receptors are often found at presynaptic terminals where they modulate calcium or potassium conductances. It has been shown that bipolar cell terminals receive a GABAergic synaptic input (Vaughn et al., 1981; Wu et al., 1981; Tachibana and Kaneko, 1987). The narrow field sustained-responding amacrine cells appear to be GABAergic (Werblin et al., 1988). This suggests that transient activity measured in wide field amacrine cells is formed at a population of bipolar cell terminals by GABAergic feedback from narrow field amacrine cells at GABAB receptors.

Animals↗
PubMed1988

A slowly inactivating potassium current truncates spike activity in ganglion cells of the tiger salamander retina.

Voltage-gated ganglion cell membrane currents were studied under whole-cell patch clamp in isolation and in retinal slices. The cells were identified by (1) backfilling their axons with rhodamine and later identifying them by their fluorescence in the slice or the mix of isolated cells or (2) by filling them with Lucifer yellow during recording in retinal slices. Both methods yielded cells with similar currents. In some cases, isolated cells lacked processes yet showed currents similar to other cells, suggesting that voltage-gated currents in all cells were located primarily at the soma. Both a conventional inactivating sodium current and a sustained calcium current were found. We describe 3 inactivating outward currents, ordered in their rate of inactivation. The fastest current resembled IA reported by Connor and Stevens (1971a, b). A slower current labeled IB inactivated with a time constant of 339 msec at 0 mV. The current with slowest inactivation is labeled IC here, inactivating with a time constant of 4.03 sec at 0 mV. An additional outward current was sustained and calcium dependent labeled IK(Ca). IB was the largest of these currents. It was slower than IA, was not blocked by 4AP, and inactivated over a much more positive potential range. IB appears to play an important role in spike generation, different from that of IA: Its inactivation leads to a slow depolarizing shift of the membrane during a current step, truncating spike activity after about 300-700 msec as the membrane potential enters the region of sodium inactivation. We analyze how the inactivating outward current acts to ensure a graded spiking response and to truncate the spiking output in the presence of large excitatory inputs.

Action Potentials↗
PubMed1987

The interaction of ionic currents mediating single spike activity in retinal amacrine cells of the tiger salamander.

We investigated the ionic interactions responsible for the characteristic nonrepetitive spike activity of amacrine cells. First we measured 4 pharmacologically separable ionic components: a voltage-gated, transient inward sodium current, a voltage-gated, sustained inward calcium current, a calcium-gated, sustained outward potassium current, and a voltage-gated, transient outward potassium current. The measurements provided the time course and magnitudes of the underlying conductances as functions of voltage. Each current was simulated following conventional Hodgkin-Huxley theory. A composite of the simulated currents was analytically reassembled to generate an approximation of the voltage response to a current step. By artificially varying the magnitude and kinetics of the different conductances in the simulation, we determined the range of values that supported the nonrepetitive spike-like response. Amacrine cells tend to remain refractory following an initial spike because (1) the entire activation range for potassium is located at positive potentials with respect to sodium inactivation, so sodium inactivation is never fully extinguished, and (2) the fully activated sodium conductance is of insufficient magnitude to subsequently reach threshold, given this residual inactivation. Shifting the sodium inactivation range by 10 mV, or increasing sodium conductance by 5 times, leads to a more repetitive form of activity. Changes in the magnitude, time course, or activation range of the potassium conductance cannot alter these conditions.

Animals↗