Wave onset in central gray matter - its intrinsic optical signal and phase transitions in extracellular polymers.

The brain is an excitable media in which excitation waves propagate at several scales of time and space. "One-dimensional" action potentials (millisecond scale) along the axon membrane, and spreading depression waves (seconds to minutes) at the three dimensions of the gray matter neuropil (complex of interacting membranes) are examples of excitation waves. In the retina, excitation waves have a prominent intrinsic optical signal (IOS). This optical signal is created by light scatter and has different components at the red and blue end of the spectrum. We could observe the wave onset in the retina, and measure the optical changes at the critical transition from quiescence to propagating wave. The results demonstrated the presence of fluctuations preceding propagation and suggested a phase transition. We have interpreted these results based on an extrapolation from Tasaki's experiments with action potentials and volume phase transitions of polymers. Thus, the scatter of red light appeared to be a volume phase transition in the extracellular matrix that was caused by the interactions between the cellular membrane cell coat and the extracellular sugar and protein complexes. If this hypothesis were correct, then forcing extracellular current flow should create a similar signal in another tissue, provided that this tissue was also transparent to light and with a similarly narrow extracellular space. This control tissue exists and it is the crystalline lens. We performed the experiments and confirmed the optical changes. Phase transitions in the extracellular polymers could be an important part of the long-range correlations found during wave propagation in central nervous tissue.

Correlation of the electrical and intrinsic optical signals in the chicken spreading depression phenomenon.

This paper presents some results on the correlation between the electrophysiological and intrinsic optical signals (IOS) of spreading depression waves in chicken retinae. We first show that the peak of the time derivative of the electrophysiological wave occurs precisely when the optical signal reaches the electrode tip. Second, by comparing bath applications of propranolol and glycerol it can be shown that the slow potential shift is not directly correlated to the intrinsic optical signal. Propranolol depresses the amplitude of the electrical wave, although the intrinsic optical signal continues being visible. On the other hand, we observe total absence of the IOS under glycerol, while the electrical wave is always present. Correlations of this kind are relevant for a deeper understanding of the underlying mechanisms of the spreading depression phenomenon.

Exogenous application of gangliosides changes the state of excitability of retinal tissue as demonstrated by retinal spreading depression experiments.

Gangliosides are amphiphilic, sialic acid-containing glycosphingolipids which are found preferentially in complex composition in the cellular membranes of the nervous system of vertebrates, including the vertebrate retina as well as in other membranes. They are always exposed to the extracellular side of the membranes. By virtue of the negative charges they carry at their headgroup, they contribute to the surface charge of the membrane and may affect ion distribution, mainly that of protons and calcium ions, at the outer side of the membranes. Using retinal spreading depression (RSD) as a tool, we show in this study that the addition of exogenous gangliosides to the extracellular space can change the state of excitability of the retinal tissue. In RSD experiments it reduces the propagation velocity as well as the intrinsic optical signal of RSD waves. These effects are concentration dependent (IC50 about 20 microM) and increase with the increasing negative charge of the ganglioside headgroup. As a possible mechanistic basis of the changes found, the change of the calcium homeostasis of the extracellular space by the exogenously added gangliosides is discussed. Gangliosides have been reported to be useful in the treatment of some neuropathological syndromes, including migraine, although experimental verification has not been possible up to now. Taking into account that the retina is a true part of the CNS, our data may be interpreted as the requested verification.

Influence of insulin-like growth factors and insulin on the [35S]sulfate uptake by cartilage of the axolotl (Ambystoma mexicanum).

The actions of mammalian insulin-like growth factors (IGF-I and IGF-II) and insulin on skeletal growth of the axolotl, Ambystoma mexicanum, were examined by monitoring the in vitro uptake of [35S]sulfate by cartilage. Both growth factors stimulated sulfate uptake significantly at a concentration of 13 nM. The increase after incubation with 130 nM insulin was similar (uptake ca. 160% of control), but the effect was not significant. Further, the binding of 125I-IGF-I and 125I-IGF-II was studied in the skeletal tissue by quantitative in vitro autoradiography. Specific binding sites for both growth factors were localized in the perichondrial tissue. 125I-IGF-II binding sites were saturated at the used doses. Scatchard and Hill plots demonstrate heterogeneous binding sites exhibiting positive cooperativity. IGF-II was more effective than IGF-I in competitively displacing both labeled ligands. Insulin only slightly affected the binding at the highest concentrations. These results indicate that the insulinlike growth factors regulate skeletal growth in urodeles as in mammals.

A depressäo alastrante sob o efeito de bário / Spreading depression under the effect of barium

Apresentamos aqui os resultados de medidas ópticas e eletro-fisiológicas obtidas durante a ocorrência de ondas de depressão alastrante (DA) na retina de galinha in vitro. Adquirindo simultaneamente os sinais supracitados foi possível analisar a sua relaçäo temporal e espacial durante a DA. As ondas foram elicitadas sob condiçöes de controle e com a aplicaçäo de cloreto de bário à soluçäao de perfusäo. Algumas das características mais afetadas foram a variaçäo lenta do potencial extra-celular, a velocidade e o valor absoluto do pico da derivada do sinal de tensäo. A onda óptica intrínseca (IOS) também foi afetada. Apesar disso, o pico da derivada (do potencial) ocorreu senpre no momento da chegada da onda óptica à ponta da micropipeta. Como o bário é um bloqueador de células gliais, os resultados dos experimentos evidenciam a relaçäo entre a despolarizaçäo glial e outros fenômenos comcomitantes tais como a onda de tensäo e o IOS, o que deverá ser objeto de investigaçöes futuras

Abstract - ln this paper we present the results of optical and electro-physiological measurements carried out during the occurrence of spreading depression waves in the in vitro chicken retina. Simultaneously acquiring the optical and electrical signals, we could verify the temporal and spatial relationship between them. The waves have been elicited under control conditions and with the application of barium chloride to the perfusing solution. Some of the most affected features of the waves were the negative slow potential shift, the velocity and the peak absolute value of the derivative of the potential wave. The intrinsic optical signal (IOS) was also affected. Nevertheless, the peak value ofthe derivative occurred always when the !OS was exactly on the tip ofthe micropipette. As longas bariurn is a K-channel blocker specific for glial cells, the results of these experiments pennit to conclude that the relationship between the glial depolarization and the concornitant phenomena such as the potential wave and the !OS exists and should be the subject offurther investigation

Regulation of interrenal secretion in the axolotl, Ambystoma mexicanum.

The regulation of corticosteroid secretion of the adrenal cortex (interrenal tissue) of axolotl (Ambystoma mexicanum) was studied using in vitro preparations of kidney containing interrenal tissue. Normally, 0.3-0.65 ng/5 min corticosterone and 0.15-0.3 ng/5 min aldosterone were released from the tissue. Regulatory peptides were effective in the following range: ACTH = arginine vasotocin > urotensin II > angiotensin II. They stimulate an elevation of corticosterone (plus 0.2-1 ng/5 min) and of aldosterone (plus 0.05-0.15 ng/5 min). The three primary effector systems leading to second messengers, adenylate cyclase (forming cAMP), phospholipase C (forming InsP3 + DAG), and phospholipase A2 (liberating arachidonic acid) are involved in stimulation of biosynthesis. It can be suggested that the second messengers stimulate the biosynthesis at the level of the steps between pregnenolone and corticosterone ('3 beta-hydroxysteroid-dehydrogenase etc.), because the release of corticosterone is more stimulated than aldosterone. This is different than the regulation of anuran interrenal tissue. Ca++ ions are involved in corticosterone secretion. Verapamil inhibits immediately the secretion of corticosteroids and elevation of external Ca++ stimulates the release. It is suggested that Ca++ mediates the secretion process itself. Metamorphosis does not change the response of the interrenal gland compared with the neotenic animal.