Histopathology of Andean Potato (Solanum tuberosum Andigenum group) varieties parasitized by the false root-knot nematode, Nacobbus aberrans / Histopatologia de variedades de batata andina (Solanum tuberosum grupo Andigenum) parasitadas pelo falso nematóide das galhas, Nacobbus aberrans

Abstract Landraces of the Solanum tuberosum Andigenum group are abundant and diverse. They are a valuable genetic resource possessing resistance to pests, diseases, and environmental stresses. In the Andean region, populations of the false root-knot nematode Nacobbus aberrans became specialized to infect native potatoes, being one of the major limiting factors affecting this crop. A better understanding of the host plant-parasite interactions is important in order to select tolerant or resistant plants to be included in management programs. Despite the close of association of N. aberrans with potato, and the great diversity of the S. tuberosum Andigenum group, few histopathological studies have been conducted. The aim of this work was to analyze histological alterations induced by different Argentine populations of the nematode in naturally infested roots of four Andean potato varieties (Collareja, Negra Imilla, Ojo de Señorita and Colorada). All the varieties showed hyperplastic tissue in the central zone of galls, where syncytia developed in close association with the nematode female. Syncytia were composed of modified hyperplastic tissue and parenchyma xylem cells. The results showed differences among varieties in their response to nematode populations, with Ojo de Señorita and Negra Imilla being the most susceptible ones. This study is the first describing histopathological alterations induced by N. aberrans in susceptible Andean potato landraces.

Resumo As variedades de Solanum tuberosum grupo Andigenum são abundantes e diversas, além de serem uma fonte importante de recursos genéticos antea resistência de pragas, doenças e estresse ambiental. Na região andina, populações de Nacobbus aberrans se especializaram em parasitar batatas nativas e representam o principal fator limitante que afeta a produção desse cultivo. Um melhor entendimento da interação hospedeiro-parasita é importante para a seleção de plantas tolerantes ou resistentes a serem incluídas nos programas de manejo. Apesar da íntima associação de N. aberrans com a batata e a grande diversidade de S. tuberosum grupo Andigenum, os estudos histopatológicos desenvolvidos são limitados. O objetivo do presente trabalho foi analisar as alterações histológicas de raízes de quatro variedades de batata andina (Collareja, Negra Imilla, Ojo de Señorita e Colorada) parasitadas naturalmente por distintas populações argentinas do nematóide. Em todas as variedades se observou tecido hiperplásico na zona central das galhas onde, também, se desenvolveram os sincícios em íntima relação com as fêmeas do nematóide. Os sincícios estavam constituídos por tecido hiperplásico modificado e por células parenquimáticas do xilema. Os resultados mostraram diferenças entre as variedades de batata frente às populações do nematóide, sendo Ojo de Señorita e Negra Imilla as que evidenciaram maior susceptibilidade. Este estudo constitui a primeira avaliação da histopalogia induzida por N. aberrans em variedades suscetíveis de batata andina.

Histopathology of Andean Potato (Solanum tuberosum Andigenum group) varieties parasitized by the false root-knot nematode, Nacobbus aberrans.

Landraces of the Solanum tuberosum Andigenum group are abundant and diverse. They are a valuable genetic resource possessing resistance to pests, diseases, and environmental stresses. In the Andean region, populations of the false root-knot nematode Nacobbus aberrans became specialized to infect native potatoes, being one of the major limiting factors affecting this crop. A better understanding of the host plant-parasite interactions is important in order to select tolerant or resistant plants to be included in management programs. Despite the close of association of N. aberrans with potato, and the great diversity of the S. tuberosum Andigenum group, few histopathological studies have been conducted. The aim of this work was to analyze histological alterations induced by different Argentine populations of the nematode in naturally infested roots of four Andean potato varieties (Collareja, Negra Imilla, Ojo de Señorita and Colorada). All the varieties showed hyperplastic tissue in the central zone of galls, where syncytia developed in close association with the nematode female. Syncytia were composed of modified hyperplastic tissue and parenchyma xylem cells. The results showed differences among varieties in their response to nematode populations, with Ojo de Señorita and Negra Imilla being the most susceptible ones. This study is the first describing histopathological alterations induced by N. aberrans in susceptible Andean potato landraces.

Histopathology of Andean Potato (Solanum tuberosum Andigenum group) varieties parasitized by the false root-knot nematode, Nacobbus aberrans

Abstract Landraces of the Solanum tuberosum Andigenum group are abundant and diverse. They are a valuable genetic resource possessing resistance to pests, diseases, and environmental stresses. In the Andean region, populations of the false root-knot nematode Nacobbus aberrans became specialized to infect native potatoes, being one of the major limiting factors affecting this crop. A better understanding of the host plant-parasite interactions is important in order to select tolerant or resistant plants to be included in management programs. Despite the close of association of N. aberrans with potato, and the great diversity of the S. tuberosum Andigenum group, few histopathological studies have been conducted. The aim of this work was to analyze histological alterations induced by different Argentine populations of the nematode in naturally infested roots of four Andean potato varieties (Collareja, Negra Imilla, Ojo de Señorita and Colorada). All the varieties showed hyperplastic tissue in the central zone of galls, where syncytia developed in close association with the nematode female. Syncytia were composed of modified hyperplastic tissue and parenchyma xylem cells. The results showed differences among varieties in their response to nematode populations, with Ojo de Señorita and Negra Imilla being the most susceptible ones. This study is the first describing histopathological alterations induced by N. aberrans in susceptible Andean potato landraces.

Resumo As variedades de Solanum tuberosum grupo Andigenum são abundantes e diversas, além de serem uma fonte importante de recursos genéticos antea resistência de pragas, doenças e estresse ambiental. Na região andina, populações de Nacobbus aberrans se especializaram em parasitar batatas nativas e representam o principal fator limitante que afeta a produção desse cultivo. Um melhor entendimento da interação hospedeiro-parasita é importante para a seleção de plantas tolerantes ou resistentes a serem incluídas nos programas de manejo. Apesar da íntima associação de N. aberrans com a batata e a grande diversidade de S. tuberosum grupo Andigenum, os estudos histopatológicos desenvolvidos são limitados. O objetivo do presente trabalho foi analisar as alterações histológicas de raízes de quatro variedades de batata andina (Collareja, Negra Imilla, Ojo de Señorita e Colorada) parasitadas naturalmente por distintas populações argentinas do nematóide. Em todas as variedades se observou tecido hiperplásico na zona central das galhas onde, também, se desenvolveram os sincícios em íntima relação com as fêmeas do nematóide. Os sincícios estavam constituídos por tecido hiperplásico modificado e por células parenquimáticas do xilema. Os resultados mostraram diferenças entre as variedades de batata frente às populações do nematóide, sendo Ojo de Señorita e Negra Imilla as que evidenciaram maior susceptibilidade. Este estudo constitui a primeira avaliação da histopalogia induzida por N. aberrans em variedades suscetíveis de batata andina.

BW284c51 blocks nicotinic acetylcholine receptors transplanted to Xenopus oocytes.

We have studied the effects of BW284c51 on the function of Torpedo nicotinic acetylcholine (Ach) receptors (nAchRs) transplanted to Xenopus laevis oocytes. BW284c51 reversible inhibited Ach-elicited currents (IAch) in a concentration-dependent manner, increased IAch desensitisation and changed the Ach concentration-dependence of the IAch from a two-site to a single-site Hill equation, without affecting the EC50. These effects were only present at hyperpolarising potentials, suggesting that nAchR blockade by BW284c51 is non-competitive and likely due to an open channel block as the principal mechanism.

Functional transplantation of chloride channels from the human syncytiotrophoblast microvillous membrane to Xenopus oocytes.

The materno-fetal transfer of metabolites and nutrients requires the operation of specific transport mechanisms through syncytiotrophoblast membranes. Electrophysiological studies on these cells are scarce and, because of their syncytial nature, whole-cell current recordings have not been carried out. We have now studied whether or not ion channels from the human syncytiotrophoblast microvillous (hSM) membrane can be transplanted to Xenopus oocytes. Sixty-two percent of hSM-injected oocytes displayed lower resting potential and higher membrane conductance than uninjected cells. The increased membrane conductance was due to the incorporation of Cl(-) channels, because neither replacing Na(+) in the bathing solution by N-methyl- D-glucamine or K(+), nor withdrawing Ca(2+) had any significant effect on the currents elicited by voltage pulses. In contrast, substitution of Cl(-) by different anions markedly affected the membrane conductance, giving an anion selectivity sequence of I(-)>Br(-)>Cl(-)>methanosulfonate congruent with gluconate. In addition, disulfonic stilbenes and gluconate, but not anthracene-9-carboxylic acid, blocked the transplanted channels. These properties are compatible with those of placental Cl(-) "maxi" channels. It is concluded that functional Cl(-) channels from the hSM become effectively incorporated into the Xenopus oocyte membrane, where their function can be studied in detail.

Ca(2+) permeability of human heteromeric nAChRs expressed by transfection in human cells.

The Ca(2+) permeability of the human heteromeric alpha 3 beta 4, alpha 4 beta 2 and alpha 4 beta 4 neuronal nicotinic acetylcholine receptors (nAChRs) was estimated by measuring the fractional Ca(2+) current (P(f)) flowing through the ligand-activated receptor-channels. Simultaneous recordings of transmembrane currents and fluorescence transients, using the whole-cell patch-clamp technique combined with fura-2 fluorescence microscopy, were performed in transiently transfected human cells. The human alpha 4 beta 2 nAChR showed a P(f) value of 2.6%, while the human alpha 3 beta 4 nAChR showed a similar P(f) value of 2.7%. Conversely, alpha 4 beta 4 nAChR exhibited a P(f) value (1.5%) significantly smaller than those of both alpha 4 beta 2 and alpha 3 beta 4 nAChRs. In test experiments performed in HEK 293 cells stably expressing rat GluR1 AMPA receptor subunit, we repeated the determination of P(f), whose value (3.2%) has previously been reported by others using the same fluorescent dye; and we found a very similar P(f) value (3.5%). In further test experiments, we found that P(f) values of chick alpha 3 beta 4 (4.4%) and alpha 4 beta 4 (2.1%) matched those previously reported by us using confocal fluorescence microscopy. Thus, our findings are consistent with those elsewhere reported even using different experimental procedures, giving a strong support to the following sequence of Ca(2+) permeability: h-alpha 3 beta 4>h-alpha 4 beta 2>h-alpha 4 beta 4.

Chemokine receptor CXCR2 regulates the functional properties of AMPA-type glutamate receptor GluR1 in HEK cells.

Experiments were conducted in both HEK cells and cerebellar neurons to investigate whether CXC chemokine receptor 2 (CXCR2) is functionally coupled to GluR1. The co-expression of CXCR2 with GluR1 in HEK cells increased (i) the GluR1 "apparent" affinity for the transmitter; (ii) the GluR1 channel open probability; and (iii) GluR1 binding site cooperativity upon CXCR2 stimulation with CXC chemokine ligand 2 (CXCL2). The affinity of C-terminal-deleted GluR1 for glutamate (Glu) remained stable instead. Furthermore, CXCL2 increased the binding site cooperativity of AMPA receptors in rat cerebellar granule cells; and the amplitude of spontaneous excitatory postsynaptic current (sEPSCs) in Purkinje neurons (PNs). Our findings indicate that the coupling of CXCR2 with GluR1 may modulate glutamatergic synaptic transmission.

Food entrainment to 4-h T cycles in rats kept under constant lighting conditions.

The effect of 4-h feeding cycles on the feeding pattern of rats kept under constant light and constant darkness, was analysed. In both cases, the scheduled pattern of food-approach behavior elicited dissociation of the feeding activity into several components associated to the feeding times (CAFT), which coexisted with the previous circadian free-running rhythms. In LL rats and in the rest phase of DD animals, the CAFT was characterized by an increased number of food approaches confined exclusively to the period of food availability, with poor anticipatory activity. However, in the active phase of some DD rats, CAFT included an additional anticipatory activity. The interaction between the CAFT and the free-running rhythms involved modulation of the CAFT, depending on the free-running periodicity. After termination of the feeding-restriction schedule, some DD and LL animals retained the CAFT for at least 3 days. However, when the access to food was blocked 10 days after ending the restricted schedule, rats did not show any feeding activity associated to the previous feeding times. A spontaneous feeding pattern similar to that imposed by the previous feeding schedule emerged immediately after food deprivation in two of the DD animals. Although these results are compatible with the existence of a food-entrainable pacemaker, the existence of a multioscillatory light-entrainable pacemaker with some oscillators entrained by food pulses and others free-running may explain our results.

Locomotor and feeding activity rhythms in a light-entrained diurnal rodent, Octodon degus.

The wheel running (WR) and feeding activity (FA) of Octodon degus, a new laboratory rodent characterized by its diurnal habits, were recorded under different lighting conditions. Under 12:12-h light-dark (LD 12:12) cycles, WR activity exhibited a crepuscular pattern with two peaks, M and E, associated with "dawn" and "dusk," respectively. In both cases, an anticipatory activity was patent, suggesting that, beside the masking effect of LD transitions, both peaks have an endogenous origin. This pattern, which was also observed under a skeleton photoperiod (LD 0.5:11.5), became unimodal after LD 0.5:23.5 and constant darkness (DD) exposure. Simultaneously, FA showed an arrhythmic pattern in most animals, especially under DD, when none of the animals exhibited a significant circadian rhythm. The existence of two groups of oscillators, or two oscillators, would explain most properties of the WR rhythms noted in this species. Our results show that the degu's temporal feeding strategy seems mainly arrhythmic, whereas its WR pattern is driven by a strongly circadian bimodal rhythm.

Food-entrained feeding and locomotor circadian rhythms in rats under different lighting conditions.

It has been suggested that two endogenous timekeeping systems, a light-entrainable pacemaker (LEP) and a food-entrainable pacemaker (FEP), control circadian rhythms. To understand the function and interaction between these two mechanisms better, we studied two behavioral circadian rhythmicities, feeding and locomotor activity, in rats exposed to two conflicting zeitgebers, food restriction and light-dark cycles. For this, the food approaches and wheel-running activity of rats kept under light-dark (LD) 12:12, constant darkness (DD), or constant light (LL) conditions and subjected to different scheduled feeding patterns were continuously recorded. To facilitate comparison of the results obtained under the different lighting conditions, the period of the feeding cycles was set in all three cases about 1h less than the light-entrained or free-running circadian rhythms. The results showed that, depending on the lighting conditions, some components of the feeding and wheel-running circadian rhythms could be entrained by food pulses, while others retained their free-running or light-entrained state. Under LD, food pulses had little influence on the light-entrained feeding and locomotor rhythms. Under DD, relative coordination between free-running and food-associated rhythms may appear. In both cases, the feeding activity associated with the food pulses could be divided into a prominent phase-dependent peak of activity within the period of food availability and another afterward. Wheel-running activity mainly followed the food pulses. Under LL conditions, the food-entrained activity consisted mainly of feeding and wheel-running anticipatory activity. The results provide new evidence that lighting conditions influence the establishment and persistence of food-entrained circadian rhythms in rats. The existence of two coupled pacemakers, LEP and FEP, or a multioscillatory LEP may both explain our experimental results.

Repeated short-fasting modifies the macronutrient self-selection pattern in rats.

The daily caloric intake and circadian pattern of macronutrient self-selection were examined in rats subjected to 3 h of food and water deprivation at the beginning or at the end of darkness. When one sole 3-h period of deprivation was applied, rats showed a compensatory response characterized by an unscheduled diurnal and nocturnal increase in the intake of the three macronutrients. However, repeated short restrictions during 15 days promoted a scheduled time-dependent feeding response, characterized by an exclusive increase in carbohydrate and fat intake and a decrease in protein intake. Repeated deprivation at the onset of dark produced a feeding response confined to the dark phase, while late dark deprivation produced both a diurnal and nocturnal increase in feeding. After 15 days of repeated restriction, rats showed no body weight variations with respect to control rats fed ad libitum. These results show that short fasting elicits a time- and macronutrient-dependent feeding response in rats, which involves reorganization of the macronutrient self-selection pattern to promote a total daily caloric compensation. These results suggest that animals principally respond to the energy deficit produced by restriction.

Feeding behavior and entrainment limits in the circadian system of the rat.

The entrainment limits of the circadian rhythms of feeding activity were studied in Wistar rats exposed to gradually increasing and decreasing or to static light-dark cycles. In the former, the entrainment limits of feeding behavior were 22 h 10 min and 26 h 40 min. In the latter, the upper limit was higher, because rats under zeitgeber period (t) length = 27 h (t27) and t28 met the criteria of entrainment. The lower limit, on the other hand, was not modified because none of the t22 animals showed entrained rhythms and one-half of the t23 rats exhibited two components in their circadian feeding rhythms, one with a period of 23 h and the other free running. This 23-h component reflected not only the masking effect of light-dark cycles but also seemed a true light-entrained component. In well-synchronized animals, food intake seemed to depend more on the number of cycles that the animal experienced than on actual time lived; however, other feeding parameters, such as meal frequency and feeding duration, remained constant when expressed per 24 h, irrespective of the t cycle. These results concerning feeding duration, meal frequency, and food intake revealed that the homeostatic and circadian controls interacted to a degree that depended on the type of variable considered. In conclusion, the entrainment limits appeared much more imprecise than they were previously thought to be, because the circadian system can only be partially synchronized near its entrainment limits. The hypothesis that the rat's circadian system is composed of multiple oscillators with different intrinsic frequencies and varying capacities for light synchronization would explain the partial desynchronization observed near the entrainment limits.

Coupling effect of locomotor activity on the rat's circadian system.

Exercise is recognized to affect circadian rhythmicity in a variety of ways. It masks the expression of other behavioral and physiological rhythms, entrains the master pacemaker, and influences the free-running period of other rhythms. In this paper we study the influence of exercise on the organization of the timing system by analyzing the effect of voluntary locomotor activity on the circadian feeding behavior of rats subjected to different lighting conditions. The availability of wheel running prevented loss of feeding circadian rhythmicity under constant bright light (LL) but did not elicit any circadian pattern in rats showing a previous arrhythmic pattern. Under dim red light (DR), the rhythm was more pronounced in exercising than in sedentary rats, while wheel-running availability accelerated the emergence of circadian rhythmicity in arrhythmic animals that were moved from LL to DR. These results can be explained by the existence of a positive feedback loop between physical exercise and its pacemaker and also suggest that exercise changes the functioning of the circadian system to facilitate the emergence of circadian rhythms in previously arrhythmic animals.

A contact eatometer suitable for feeding restriction schedules.

We describe and test a programmable feeding system based on a contact eatometer, previously developed in our laboratory, consisting of a swinging grid beneath a hopper that is moved by the rat each time it wants to eat. This has not been connected to a solenoid that can block the movement of the grid to prevent access to the food. It also provides a continuous record of food approaches even during restricted periods when no food is made available, and so it is useful in studying the synchronizing role of food in the circadian rhythms of feeding behavior. The simplicity and cheapness of its construction and the effective blocking of access to food make it an ideal tool in chronobiological studies involving the simultaneous use over long periods of a large number of animals.

The zero dispersion limit for the Korteweg-deVries KdV equation.

We use the inverse scattering method to determine the weak limit of solutions of the Korteweg-deVries equation as dispersion tends to zero. The limit, valid for all time, is characterized in terms of a quadratic programming problem which can be solved with the aid of function theoretic methods. For large t, the solutions satisfy Whitham's averaged equations at some times and the equations found by Flaschka et al. at other times.

Separation and quantification of urinary di- and polyamines by gas chromatography with electron capture detection.

A sensitive and specific gas chromatographic assay procedure employing electron capture detection has been developed for the assay of free and total di- and polyamines in human urine. Urine samples, hydrolysed with hydrochloric acid where necessary for the measurement of total amine output, were evaporated to dryness and, after the residues had been taken up in water, purified successively on Porapak Q and Dowex 50 X2 columns. Following evaporation of eluate, pentafluoropropionyl derivatives were made and analysed gas chromatographically using temperature programming. Di- and polyamines can be measured accurately at the picomole level and normal urinary output values calculated using this method agree well with those noted by other workers.