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PubMedDate not supplied

Estimating the contributions of NMDA and non-NMDA currents to EPSPs in retinal ganglion cells.

Whole-cell recordings were obtained from retinal ganglion cells of the tiger salamander (Ambystoma tigrinum) in a superfused slice preparation to evaluate contributions of NMDA (N-methyl-D-aspartate) and KA/AMPA (kainate/alpha-amino-3-hydroxy-5-methyl-4-isoxalone propionic acid) receptors to excitatory postsynaptic potentials (EPSPs) of retinal ganglion cells. Synaptic activation of retinal ganglion cells was achieved through the use of a brief pressure pulse of hyperosmotic Ringer (Ringer + sucrose) delivered through a microelectrode visually placed in the inner plexiform layer while whole-cell recordings were obtained from adjacent cells in the ganglion cell layer. Separation of NMDA and KA/AMPA excitatory postsynaptic currents (EPSCs) was achieved through the application of the antagonists NBQX and D-AP7, while inhibitory currents were blocked by strychnine and picrotoxin. Simple addition of the two independent EPSCs showed, most often, that the sum of the KA/AMPA and NMDA currents was less than the control response, but in some cases the sum of the two currents exceeded the magnitude of the control response. Neither result was consistent with expectations based on voltage-clamp principles and the assumption that the two currents were independent; for this reason, we considered the possibility of nonlinear interactions between KA/AMPA and NMDA receptors. Computer simulations were carried out to evaluate the summation experiments. We used both an equivalent cylinder model and a more realistic, compartmental model of a ganglion cell constrained by a passive leakage conductance, a linear KA/AMPA synaptic current, and a nonlinear NMDA current based on the well-known, voltage-sensitive Mg2+ block. Computer simulation studies suggest that the hypo- and hyper-summation of NMDA and KA/AMPA currents, observed physiologically, can be accounted for by a failure to adequately space clamp the neuron. Clamp failure leads to enhanced NMDA currents as the ion channels are relieved of the Mg2+ block; their contribution is thus exaggerated depending on the magnitude of the conductance change and the spatial location of the synaptic input.

2-Amino-5-phosphonovalerate↗
PubMedDate not supplied

Chloride efflux inhibits single calcium channel open probability in vertebrate photoreceptors: chloride imaging and cell-attached patch-clamp recordings.

The present study uses cell-attached patch-recording techniques to study the single-channel properties of Ca2+ channels in isolated salamander photoreceptors and investigate their sensitivity to reductions in intracellular Cl-. The results show that photoreceptor Ca2+ channels possess properties similar to L-type Ca2+ channels in other preparations, including (1) enhancement of openings by the dihydropyridine agonist, (-)BayK8644; (2) suppression by a dihydropyridine antagonist, nisoldipine; (3) single-channel conductance of 22 pS with 82 mM Ba2+ as the charge carrier; (4) mean open probability of 0.1; (5) open-time distribution fit with a single exponential (tau0 = 1.1 ms) consistent with a single open state; and (6) closed time distribution fit with two exponentials (tau(c1) = 0.7 ms, tau(c2) = 25.4 ms) consistent with at least two closed states. Using a Cl- -sensitive dye to measure intracellular [Cl-], it was found that perfusion with gluconate-containing, low Cl- medium depleted intracellular [Cl-]. It was therefore possible to reduce intracellular [Cl-] by perfusion with a low Cl- solution while maintaining the extracellular channel surface in high Cl- pipette solution. Under these conditions, the single-channel conductance was unchanged, but the mean open probability fell to 0.03. This reduction can account for the 66% reduction in whole-cell Ca2+ currents produced by perfusion with low Cl- solutions. Examination of the open and closed time distributions suggests that the reduction in open probability arises from increases in closed-state dwell times. Changes in intracellular [Cl-] may thus modulate photoreceptor Ca2+ channels.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMedDate not supplied

Computational analysis of vertebrate phototransduction: combined quantitative and qualitative modeling of dark- and light-adapted responses in amphibian rods.

We evaluated the generality of two models of vertebrate phototransduction. The approach was to quantitatively optimize each model to the full waveform of high-quality, dark-adapted (DA), salamander rod flash responses. With the optimal parameters, each model was then used to account for signature, qualitative features of rod responses from three experimental paradigms (stimulus/response, "S/R suite"): (1) step responses; (2) the intensity dependence of the period of photocurrent saturation (Tsat vs. ln(I)); and (3) light-adapted (LA) incremental flash sensitivity as a function of background intensity. The first model was the recent successful model of Nikonov et al. (1998). The second model replaced the instantaneous Ca2+ buffering used in the Nikonov et al. model with a dynamic buffer. The results showed that, in the absence of the dynamic Ca2+ buffer, the Nikonov et al. model does not have sufficient flexibility to provide a good fit to the flash responses, and, using the same parameters, reproduce the salient features of the S/R suite--critical features at step onset and offset are absent; the Tsat function has too shallow a slope; and the model cannot generate the empirically observed I-range of Weber-Fechner LA behavior. Some features could be recovered by changing parameters, but only at the expense of the fit to the reference (Ref) data. When the dynamic buffer is added, the model is able to achieve an acceptable fit to the Ref data while reproducing several features of the S/R suite, including an empirically observed Tsat function, and an extended range of LA flash sensitivity adhering to Weber's law. The overall improved behavior of the model with a dynamic Ca2+ buffer indicates that it is an important mechanism to include in a working model of phototransduction, and that, despite the slow kinetics of amphibian rods, Ca2+ buffering should not be simulated as an instantaneous process. However, neither model was able to capture all the features with the same parameters yielding the optimal fit to the Ref data. In addition, neither model could maintain a good fit to the Ref data when five key biochemical parameters were held at their current known values. Moreover, even after optimization, a number of important parameters remained outside their empirical estimates. We conclude that other mechanisms will need to be added, including additional Ca2+-feedback mechanisms. The present research illustrates the importance of a hybrid qualitative/quantitative approach to model development, and the limitations of modeling restricted sets of data.

Adaptation, Ocular↗
PubMedDate not supplied

Amacrine and ganglion cell contributions to the electroretinogram in amphibian retina.

The neuronal generators of the b- and d-waves of the electroretinogram (ERG) were investigated in the tiger salamander retina to determine if amacrine and ganglion cells contribute to this field potential. Several agents were used that affect third-order neurons, such as tetrodotoxin, baclofen, and NMDA agonists and antagonists. Baclofen, an agent that enhances light responses in third-order neurons, increased the d-wave and reduced the b-wave. In contrast, agents that decrease light responses in third-order neurons had the opposite effect of enhancing the b-wave and depressing the d-wave. The effect on the d-wave was particularly pronounced. The results indicate that third-order neuronal activity influences b- and d-waves of the ERG. The opposing actions suggest that the b-wave to d-wave ratio might serve as an measure of ganglion cell function.

Ambystoma↗
PubMedDate not supplied

Effects of light adaptation on contrast processing in bipolar cells in the retina.

Effects of light adaptation on contrast processing in the outer retina were investigated over nearly four decades of background illumination by analyzing the intracellular responses of 111 bipolar cells, 66 horizontal cells, and 22 cone photoreceptors in the superfused eyecup of the tiger salamander (Ambystoma tigrinum). Light adaptation had striking and similar effects on the average contrast responses of the hyperpolarizing (Bh) and depolarizing (Bd) classes of bipolar cells: Over the lower two decades of background illumination, the contrast gain increased 7-fold to reach values as high as 20-30, the dynamic range and the half-maximum contrast decreased by about 60%, the total voltage range increased some 40%, and contrast dominance changed from highly positive to more balanced. At higher levels of background, most aspects of the contrast response stabilized and Weber's Law then held closely. In this background range, the contrast gain of bipolar cells was amplified some 20X relative to that of cones whereas the corresponding amplification in horizontal cells was about 6X. Differences in the growth of contrast gain with the intensity of the background illumination for cones versus bipolar cells suggest that there are at least two adaptation-dependent mechanisms regulating contrast gain. One is evident in the cone photoresponse such that an approximately linear relation holds between the steady-state hyperpolarization and contrast gain. The other arises between the voltage responses of the cones and bipolar cells. It could be presynaptic (modulation of cone transmitter release by horizontal cell feedback or other mechanisms) and/or postsynaptic, that is, intrinsic to bipolar cells. Contrast gain grew with the background intensity by a larger factor in horizontal than in bipolar cells. This provides a basis for the widely held view that light adaptation increases the strength of surround antagonism in bipolar cells. On average, the effects of light adaptation and most quantitative indices of contrast processing were remarkably similar for Bd and Bh cells, implying that both classes of bipolar cells, despite possible differences in underlying mechanisms, are about equally capable of encoding all primary aspects of contrast at all levels of light adaptation.

Adaptation, Ocular↗
PubMedDate not supplied

Center-surround organization in bipolar cells: symmetry for opposing contrasts.

Intracellular recordings were obtained from 73 cone-driven bipolar cells in the light-adapted retina of the tiger salamander (Ambystoma tigrinum). Responses to flashes of negative and positive contrast for centered spots and concentric annuli of optimum spatial dimensions were analyzed as a function of contrast magnitude. For both depolarizing and hyperpolarizing bipolar cells, it was found that remarkably similar responses were observed for the center and surround when comparisons were made between responses of the same response polarity and thus, responses to opposite contrast polarity. Thus, spatial information and contrast polarity appear to be rather strongly confounded in many bipolar cells. As a rule, the form of the contrast/response curves for center and surround approximated mirror images of each other. Contrast gain and C50 (the contrast required for half-maximal response) were quantitatively similar for center and surround when comparisons were made for responses of the same response polarity. The average contrast gain of the bipolar cell surround was 3-5 times higher than that measured for horizontal cells. Contrast/latency measurements and interactions between flashed spots and annuli showed that the surround response is delayed by 20-80 ms with respect to that of the receptive-field center. Cones showed no evidence for center-surround antagonism while for bipolar cells, the average strength of the surround ranged from about 50% to 155% of the center, depending on the test and response polarity. The results of experiments on the effects of APB (100 microM) on depolarizing bipolar cells suggest that the relative contribution of the feedback pathway (horizontal cell to cones) and the feedforward pathway (horizontal cell to bipolar cell) to the bipolar surround varies in a distributed manner across the bipolar cell population.

Adaptation, Ocular↗
PubMedDate not supplied

Contrast encoding in retinal bipolar cells: current vs. voltage.

To investigate the influence of voltage-sensitive conductances in shaping light-evoked responses of retinal bipolar cells, whole-cell recordings were made in the slice preparation of the tiger salamander, Ambystoma tigrinum. To study contrast encoding, the retina was stimulated with 0.5-s steps of negative and positive contrasts of variable magnitude. In the main, responses recorded under voltage- and current-clamp modes were remarkably similar. In general agreement with past results in the intact retina, the contrast/response curves were relatively steep for small contrasts, thus showing high contrast gain; the dynamic range was narrow, and responses tended to saturate at relatively small contrasts. For ON and OFF cells, linear regression analysis showed that the current response accounted for 83-93% of the variance of the voltage response. Analysis of specific parameters of the contrast/response curve showed that contrast gain was marginally higher for voltage than current in three of four cases, while no significant differences were found for half-maximal contrast (C50), dynamic range, or contrast dominance. In sum, the overall similarity between current and voltage responses indicates that voltage-sensitive conductances do not play a major role in determining the shape of the bipolar cell's contrast response in the light-adapted retina. The salient characteristics of the contrast response of bipolars apparently arise between the level of the cone voltage response and the postsynaptic current of bipolar cells, via the transformation between cone voltage and transmitter release and/or via the interaction between the neurotransmitter glutamate and its postsynaptic receptors on bipolar cells.

Animals↗
PubMedDate not supplied

N-methyl-D-aspartate receptors contribute to the baseline noise of retinal ganglion cells.

Whole-cell recordings of tiger salamander ganglion cells were obtained using a superfused retinal slice preparation. Membrane current fluctuations were recorded under voltage-clamp conditions with cells usually held at -70 mV. Current fluctuations at rest (Mg2+ = 1 mM) were reduced by adding D-2-amino-7-phosphonoheptanoate (D-AP7). Resting fluctuations were increased by adding N-methyl-D-aspartate (NMDA) or by removing extracellular Mg2+. These increased fluctuations were blocked by D-AP7. Blocking NMDA receptors under control conditions also reduced a tonic inward current by -1 to -15 pA. Fluctuation analysis of current noise shows that the noise power spectrum measured in the presence of NMDA is similar to that measured under Mg(2+)-free conditions. We conclude that NMDA receptors are active in cells held at -70 mV even in the presence of 1 mM Mg2+. We believe this activation is due to the presence of endogenous glutamate in the retina. The observations of this study strongly suggest that NMDA receptors contribute to the resting noise and conductance properties of retinal ganglion cells. Our results suggest NMDA receptors are activated by an ambient level of extracellular glutamate whose source has yet to be determined.

Ambystoma↗
PubMedDate not supplied

Activation of glutamate transporters in rods inhibits presynaptic calcium currents.

We found that L-glutamate (L-Glu) inhibits L-type Ca2+ currents (ICa) in rod photoreceptors. This inhibition was studied in isolated rods or rods in retinal slices from tiger salamander using perforated patch whole cell recordings and Cl(-)-imaging techniques. Application of L-Glu inhibited ICa by approximately 20% at 0.1 mM and approximately 35% at 1 mM. L-Glu also produced an inward current that reversed around ECl. The metabotropic glutamate receptor (mGluR) agonists t-ADA (Group I), DCG-IV (Group II), and L-AP4 (Group III) had no effect on ICa. However, the glutamate transport inhibitor, TBOA (0.1 mM), prevented L-Glu from inhibiting ICa. D-aspartate (D-Asp), a glutamate transporter substrate, also inhibited ICa with significantly more inhibition at 1 mM than 0.1 mM. Using Cl imaging, L-Glu (0.1-1 mM) and D-Asp (0.1-1 mM) were found to stimulate a Cl- efflux from terminals of isolated rods whereas the ionotropic glutamate receptor agonists NMDA, AMPA, and kainate and the mGluR agonist, 1S,3R-ACPD, did not. Glutamate-evoked Cl- effluxes were blocked by the glutamate transport inhibitors TBOA and DHKA. Cl- efflux inhibits Ca2+ channel activity in rod terminals (Thoreson et al. (2000), Visual Neuroscience 17, 197). Consistent with the possibility that glutamate-evoked Cl- efflux may play a role in the inhibition, reducing intraterminal Cl- prevented L-Glu from inhibiting ICa. In summary, the results indicate that activation of glutamate transporters inhibits ICa in rods possibly as a consequence of Cl- efflux. The neurotransmitter L-Glu released from rod terminals might thus provide a negative feedback signal to inhibit further L-Glu release.

Amino Acid Transport System X-AG↗
PubMedDate not supplied

Retinal bipolar cells: contrast encoding for sinusoidal modulation and steps of luminance contrast.

Contrast encoding for sinusoidal modulations of luminance contrast was investigated by intracellular recording in the intact salamander retina. In what appears to be the first study of this kind for vertebrate bipolar cells, responses of the central receptive-field mechanism of cone-driven cells to modulation of 3 Hz were analyzed quantitatively via both signal averaging and a Fast Fourier Transform (FFT) while the retina was light adapted to 20 cd/m2. Depolarizing and hyperpolarizing bipolar cells showed very similar encoding. Both responded with sinusoidal waveforms whose amplitude varied linearly with modulation depths ranging up to 7-8%. The slope of the modulation/response curve was very steep in this range. Thus, the contrast gain was high, reaching values of 6-7, and the half-maximal response was achieved at modulations of 9% or less. At modulations above approximately 15%, the responses typically showed strong compressive nonlinearity and the waveform was increasingly distorted. At maximum modulation, the higher harmonics of the FFT constituted about 30% of the amplitude of the fundamental. Measurements were also made for cones and horizontal cells. Both cell types showed predominantly linear responses and low contrast gain, in marked contrast to bipolar cells. These results suggest that the high contrast gain and strong nonlinearity of bipolar cells largely arise postsynaptic to cone transmitter release. Further experiments were performed to compare responses to contrast steps versus those to sinusoidal modulation. In the linear range, we show that the contrast gains of cones and horizontal cells are low and virtually identical for both steps and sinusoidal modulations. In bipolar cells, on the other hand, the contrast gain is about two times greater for steps than that for the 3-Hz sine waves. These results suggest that mechanisms intrinsic to bipolar cells act like a high-pass filter with a short time constant to selectively emphasize contrast transients over slower changes in contrast.

Ambystoma↗
PubMedDate not supplied

GABA sensitivity of spectrally classified horizontal cells in goldfish retina.

We studied the GABA sensitivity of horizontal cells in the isolated goldfish retina. After the glutamatergic input to the horizontal cells was blocked with DNQX, GABA depolarized the monophasic and biphasic horizontal cells. The pharmacology of these GABA-induced depolarizations was tested with the GABA receptor antagonists bicuculline-methiodide and picrotoxin, the GABA transporter agonist nipecotic acid, and the GABA transporter antagonist SKF 89976-A. The GABA-induced responses of monophasic horizontal cells consisted of two components; one with characteristics of GABA-gated chloride channels, and one with characteristics of GABA transporters. In biphasic horizontal cells, we only found evidence for GABA-gated chloride channels. The results show that monophasic horizontal cells in goldfish contain the two components of a positive feedback loop (GABA transporters and GABA-gated chloride channels), as described in salamander. Furthermore, our results indicate that the monophasic horizontal cells may project directly to the biphasic horizontal cells, via an excitatory GABAergic pathway. We propose that the function of these GABAergic systems in horizontal cells is to abolish cone dominance in bipolar cells surround in the dark-adapted retina.

6-Cyano-7-nitroquinoxaline-2,3-dione↗
PubMedDate not supplied

Regulation of cGMP-dependent current in On bipolar cells by calcium/calmodulin-dependent kinase.

The metabotropic receptor mGluR6 is localized to the dendrites of On bipolar cells and mediates synaptic input from photoreceptors. The binding of glutamate to the receptor activates a phosphodiesterase (PDE), which then hydrolyzes cGMP. A nonselective cationic conductance, believed to be gated directly by cGMP, is turned off as a result of the fall in cGMP levels, and the cell hyperpolarizes. Here we present evidence for regulation of the conductance by an additional mechanism that it is independent of cGMP. Whole-cell recordings were obtained from On bipolar cells in slices of tiger salamander retina. Dialysis of cells with 1 microM KN-62 or 10 microM KN-93, two inhibitors of type II calmodulin-dependent protein kinase (CaMKII), depressed cGMP-dependent currents. This depression persisted when hydrolysis of cGMP was prevented with IBMX, a broad-spectrum PDE inhibitor, suggesting that CaMKII acts downstream from the PDE in the cascade. The depression of cGMP-dependent currents was probably not due to a direct interaction of the inhibitors with the channels as neither 1 microM KN-62 or 10 microM KN-93 was found to have any effect on cyclic nucleotide-gated channels when applied directly to excised patches of rod outer segments. We propose that phosphorylation by CaMKII may be an important mechanism for regulating the cGMP-dependent conductance of On bipolar cells.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMedDate not supplied

AMPA receptor kinetics limit retinal amacrine cell excitatory synaptic responses.

Amacrine cells that respond transiently to maintained illumination are thought to mediate transient inhibitory input to ganglion cells. The excitation of these transient amacrine cells is thought to be limited by inhibitory feedback to bipolar cells. We investigated the possibility that desensitizing AMPA and/or kainate (KA) receptors on amacrine cells might also limit the duration of amacrine cell excitation. To determine how these receptors might affect amacrine cell input and output, we made whole-cell recordings from amacrine and ganglion cells in the salamander retinal slice. The specific AMPA receptor antagonist GYKI-53655 blocked non-NMDA receptor-mediated amacrine cell excitatory postsynaptic currents (EPSCs) and kainate puff-elicited currents, indicating that AMPA, and not KA, receptors mediated the responses. Cyclothiazide, an agent that reduces AMPA receptor desensitization, increased the amplitude and duration of amacrine cell EPSCs. To measure the output of transient amacrine cells, we recorded glycinergic inhibitory postsynaptic currents (IPSCs) from ganglion cells, and found that these were also enhanced by cyclothiazide. Thus, prolongation of amacrine cell AMPA receptor activation enhanced amacrine cell output. Current responses elicited by puffing glycine onto ganglion cell dendrites were not affected by cyclothiazide, indicating that the enhancement of glycinergic IPSCs was not due to a direct effect on glycine receptors. These data suggest that rapid AMPA receptor desensitization and/or deactivation limits glycinergic amacrine cell excitation and the resulting inhibitory synaptic output.

Animals↗
PubMed2004

Monitoring neurotransmitter release from isolated retinas using online microdialysis-capillary electrophoresis.

Release of neurotransmitters and other primary amine-containing analytes from intact, isolated larval salamander (Ambystoma tigrinum) retinas maintained in a 6.5-microL perfusion chamber was monitored using online microdialysis-capillary electrophoresis with laser-induced fluorescence detection (CE-LIF). Primary amines were derivatized online with o-phthaldialdehyde (OPA) and beta-mercaptoethanol. With the use of overlapping injections, the perfusate was sampled every approximately 10 s. Although separation conditions were optimized using 20 mM hydroxypropyl-gamma-cyclodextrin (HP-gamma-CD) for a number of important neuromessengers including D- and L-serine, D- and L-asparate, glutamate, GABA, serotonin, dopamine, norepinephrine, and taurine, only glutamate (0.48 +/- 0.27 microM), GABA (0.25 +/- 0.12 microM), taurine (5.5 +/- 2.1 microM), and l-serine (2.8 +/- 1.0 microM) were identified in the perfusate. Elevated levels of glutamate, GABA, and taurine were detected during stimulation with 60 mM K+. This method is the first to directly sample multiple neurotransmitters from perfused, isolated retinas and to observe changes in efflux of these neurotransmitters as a result of pharmacological stimulation.

Ambystoma↗
PubMed1993

Activation of retinal rod cGMP-gated channels: what makes for an effective 8-substituted derivative of cGMP?

Analogs of cGMP bearing diverse substituents at the C8 position of the guanine ring system have been shown to activate the cGMP-activated channel of retinal rods at concentrations lower than cGMP itself. In an effort to understand this behavior, we synthesized eight novel C8-substituted derivatives and tested their ability to activate channels in excised patches from salamander rod outer segments. We express the effectiveness of each analog as a ratio (in brackets) of the concentration required to open half of the channels in a patch to that required of 8-Br-cGMP, previously shown to be about 10 times more effective than cGMP. Five of the derivatives contained a thio substitution at C8: n-propylthio-cGMP [0.61], sulfoethylthio-cGMP [0.90], carboxyethylthio-cGMP [0.97], aminoethylthio-cGMP [2.8], and (trimethylamino)ethylthio-cGMP [8.5]. Three of the derivatives contained an amino substitution at C8: carboxyethylamino-cGMP [22], n-propylamino-cGMP [25], and aminoethylamino-cGMP [230]. The results indicate that thio-substitution at C8 produces more effective analogs than does amino-substitution, regardless of the chemical nature of the terminal functional group. Derivatives containing neutral and apolar tails opened channels at much lower concentrations than their positively-charged counterparts with the same C8 substituent. Analogs having negatively-charged tails were also more effective than those with positive charge but not quite as effective as those with neutral tails.(ABSTRACT TRUNCATED AT 250 WORDS)

Ambystoma↗
PubMed2004

Role of the 9-methyl group of retinal in cone visual pigments.

In rhodopsin, the 9-methyl group of retinal has previously been identified as being critical in linking the ligand isomerization with the subsequent protein conformational changes that result in the activation of its G protein, transducin. Here, we report studies on the role of this methyl group in the salamander rod and cone pigments. Pigments were generated by combining proteins expressed in COS cells with 11-cis 9-demethyl retinal, where the 9-methyl group on the polyene chain has been deleted. The absorption spectra of all pigments were blue-shifted. The red cone and blue cone/green rod pigments were unstable to hydroxylamine; whereas, the rhodopsin and UV cone pigments were stable. The lack of the 9-methyl group of the chromophore did not affect the ability of the red cone and blue cone/green rod pigments to activate transducin. On the other hand, with the rhodopsin and UV cone pigments, activation was diminished. Interestingly, the red cone pigment containing the retinal analogue remained active longer than the native pigment. Thus, the 9-methyl group of retinal is not important in the activation pathway of the red cone and blue cone/green rod pigments. However, for the red cone pigment, the 9-methyl group of retinal appears to be critical in the deactivation pathway.

Ambystoma↗
PubMed2005

Alkaloids from amphibian skin: a tabulation of over eight-hundred compounds.

A diverse array of biologically active, lipid-soluble alkaloids have been discovered in amphibian skin. Such alkaloids include the following: the steroidal samandarines from salamanders, the batrachotoxins, histrionicotoxins, gephyrotoxins, and epibatidine from neotropical poison frogs (Dendrobatidae), the pumiliotoxins, allopumiliotoxins, homopumiliotoxins, and decahydroquinolines from certain genera of anurans from four families (Dendrobatidae, Mantellidae, Bufonidae, and Myobatrachidae), a variety of izidines (pyrrolizidines, indolizidines, quinolizidines, lehmizidines), pyrrolidines, piperidines, various tricyclics (related in structures to the coccinellines), and spiropyrrolizidines from the first three of these four families, the pseudophrynamines from one genus of Australian frogs, and a variety of unclassified alkaloids as yet of undetermined structure. With the exception of the samandarines and the pseudophrynamines, all alkaloids appear to be derived from dietary sources. Although only a few of the over 800 amphibian skin alkaloids have been detected in arthropods, putative arthropod sources for the batrachotoxins and coccinelline-like tricyclics (beetles), the pumiliotoxins (ants, mites), the decahydroquinolines, izidines, pyrrolidines, and piperidines (ants), and the spiropyrrolizidines (millipedes) have been discovered. Ants are likely sources for histrionicotoxins, lehmizidines, and tricyclic gephyrotoxins. Epibatidines represent an important alkaloid class without a putative dietary source. The structures for many of these alkaloids have been rigorously established, while the structures of others represent tentative proposals, based only on mass spectral and FTIR spectral data, along with analogies to structures of well-defined alkaloids.

Alkaloids↗
PubMed2003

Biogenesis of melanosomes in Kupffer cells of Proteus anguinus (Urodela, Amphibia).

The ultrastructural characteristics of melanosomes and premelanosomes observed during the biogenesis of melanosomes in liver pigment cells of the neotenic cave salamander Proteus anguinus (Proteidae) are described. It is well known that amphibian liver pigment cells, also known as Kupffer cells (KC), contain melanosomes and are able to synthesize melanin. Liver pigment cells of P. anguinus contain numerous siderosomes and melanosomes. The melanosomes are grouped together within single-membrane-bounded bodies, named as 'clusters of melanosomes' or 'melanosomogenesis centers'. Inside such clusters, different structures are present: (1) filament-like structures, characteristic of the initial stage of melanosome biogenesis, (2) medium electron-dense melanosomes in different stages of melanization, (3) melanosomes with an electron-dense cortical area and a less electron-dense medullar area, and (4) uniformly highly electron-dense mature melanosomes or melanin granules. Histochemical and cytochemical dihydroxyphenylalanine (DOPA) oxidase reactions in pigment cells were positive. Our results confirm the ability of amphibian KC to synthesize melanin and contribute to this little known subject.

Animals↗