Showing papers in "Journal of Neurobiology in 1973"

Multiple forms of acetylcholinesterase and their distribution in endplate and non-endplate regions of rat diaphragm muscle.

Abstract: The properties of the cholinesterase activity in homogenates of whole rat diaphragm and of innervated (+EP) and non-innervated (−EP) regions of the muscle have been investigated. Under standard assay conditions, over 90% of the cholinesterase activity of whole muscle homogenates was due to specific acetylcholinesterases. The specific activity of acetylcholinesterase was higher in +EP regions of muscle than in −EP regions. About 40% of the total activity was calculated to be specifically associated with the endplates. When a high speed supernatant fraction of muscles homogenized in 1 M NaCl, 0.5% Triton X-100 was subjected to velocity sedimentation in a sucrose gradient, and three species of acetylcholinesterase activity with sedimentation constants of 4 S, 10 S and 16 S were observed. All three forms were stable under the conditions of sedimentation and had buoyant densities of approximately 1.28. All three hydrolyzed β-methylacetylcholine at approximately 30% the rate that acetylcholine was hydrolyzed. The 10 S and 16 S forms were inhibited by concentrations of acetylcholine over 1.25 mM, but no substrate inhibition was observed with the 4 S enzyme. Velocity sedimentation of extracts from +EP and −EP regions of muscle demonstrated that the 4 S and 10 S forms of the enzyme were distributed throughout the muscle while the 16 S form was found only in +EP regions. Extracts of the phrenic nerve contained only 4 S and 10 S forms. Thus, the 16 S form of acetylcholinesterase is specifically associated with endplate regions of muscle and may correspond to the endplate enzyme. Seven days after denervation of the diaphragm, both endplate-specific cholinesterase activity and the cholinesterase activity in −EP regions of muscle were decreased. Although the activity of all three forms of acetylcholinesterase were decreased in denervated muscle, the largest proportional decrease occurred in the activity of the 16 S form.

Neural mechanisms underlying behavior in the locust Schistocerca gregaria I. Physiology of identified motorneurons in the metathoracic ganglion

Abstract: A preparation of the desert locust, Schistocera gregaria, has been developed, in which it was possible to work with identified neurons while still allowing some behavior. A total of 26 motorneurons to the hind leg were studied singly, and in various pairs, both by direct stimulation, and by recording during spontaneous activity and various reflex actions. Motorneurons were identified by passing current into their somata and correlating the evoked somata spikes with extracellularly or intracellularly recorded events in the muscles. Tension of the muscle was also recorded and motor axons were stimulated to evoke antidromic spikes in the somata. Both epsp's and ipsp's can be seen clearly in recordings from the somata; spikes appear as electrotonically conducted remnants only. Somata exhibited little or no electrogenesis. It is inferred that impulses are initiated in a zone tentatively identified with the region of emergence of the motor axon from the neuropil. Integration occurs in the neuropilar segment, with the soma serving as a parallel RC element. Data was obtained on the central mechanisms of coordination of synergistic and antagonistic motorneurons and on the modes of excitation of slow and fast neurons to the same muscles.

The neuronal basis of behavior in Tritonia. 3. Neuronal mechanism of a fixed action pattern.

Abstract: We have investigated the roles played by numerous identified brain cells in initiating and controlling the coordinated sequence of movements of an instinctive escape-swimming sequence in an intact animal preparation of the nudibranch mollusc Tritonia diomedia. Intracellular electrical activity in different neurons has been correlated with the various phases of the behavior. We recognized four major stages in the response: (1) reflex local withdrawal; (2) preparation for swimming; (3) swimming; and (4) termination. We have located and studied brain cells whose activity is associated with the following aspects of swimming: withdrawal; elongation; triggering behavior; dorsal flexion; ventral flexion; and neurons which excite both dorsal and ventral flexor neurons simulataneously. We find that specific neurons play clearly defined and invariant roles in control of escape-swimming and that the neuronal circuitry underlying the coordination of the sequence is the same in different individuals of the species. Details of the neuronal circuitry and a number of the general functional attributes of interacting cell groups have been determined directly or inferred from observations of cell to cell interactions. A preliminary model of the neuronal apparatus which controls this behavior is discussed. The principal findings are: (1) a discrete group of electrically coupled neurons determines, by its output, whether or not escapeswimming will be executed; (2) the neuronal elements responsible for execution of the swimming stages of the sequence are maintained in an excited state for the required period, in part by a regenerative feedback system; (3) alternating bursts of impulses in functional antagonists are co-ordinated in part by reciprocal inhibition between them; and (4) termination of the sequence occurs abruptly at a particular phase in the swimming cycle and appears to be an active neural process, rather than a simple running-down.

Neural mechanism underlying behavior in the locust Schistocerca gregaria. 3. Topography of limb motorneurons in the metathoracic ganglion.

Abstract: Somata of 26 of the motorneurons situated on the right side of the metathoracis ganglion of the locust S. gregaria were identified by correlating their electrical activity with extracellularly or intracellularly recorded muscle potentials evoked either reflexly or by direct elctrical stimulation of the somata through the recording microelectrode. The neurons identified included most of those innervating the major leg muscles. Each neuron occupies a relatively fixed site on the ventral surface of the ganglion. The identified neurons were filled with procion yellow, the ganglia fixed and photographed as whole mounts, and then serially sectioned to determine the three dimensional topography of the major dendrites and the pathway of the axons through the neuropil. The topographical features of individual motorneurons were sufficiently characteristic to permit identification, Nevertheless, it was not found possible to classify the neurons on the basis of branching pattern. There is no relation between location in the ganglion and the muscle innervated, but locomotor motorneurons are clustered togethers in anterior, ventral, and lateral pockets.

The neuronal basis of behavior in Tritonia. I. Functional organization of the central nervous system.

Abstract: An account is presented of the brain (paired cerebral, pleural, and pedal ganglia) of the nudibranch mollusc Tritonia diomedia. The major efferent nerve fibers are related to their nerve cell bodies and their functional roles identified as far as possible. The channels of sesory input relating to some of these neurons are given so as to provide an overall view of the organization of the brain. A standardized system of abbreviation and notation for the central ganglia, nerve trunks, and gaint somata is proposed. The system of references is intended to provide a guide to the location in the ganglia of many of the smaller neurons of which the functional attributes are known, but which cannot be consistently recognized on visual criteria alone. A system of rectangular coordinates is proposed which is applied to the natural outline of the ganglia. In addition, a system of cell notation is described which is independent of the co-ordinates used to define the position of the cell on the grid. Cell which by reason of their size, pigmentation, characteristic location and physiological attributes are consistently recognizable from animal to animal are numbered. Two principles were followed in numbering cells; (i) the series begins at unity in each ganglion; (ii) cell homologues in opposite ganglia are given the same number, but distinguished by prefixing the abbreviation for the ganglion in which they occur. It is considered that the system will facilitate the exchange of information between workers on the same species, and also benefit the comparison of neural organization of behavior in closely related forms. The brain is organized in an almost exactly bilaterally symmetrical manner. There are a few bilateral neural pathways, but the major functional routes are ipsilateral. A few motorneurons, which are uniquely identifiable anatomically, cause unique, discrete movements. Others are in small groups sharing overlapping or similar functions.

Neuronal circuit reorganization in mammalian agranular cerebellar cortex.

Abstract: Morphological and electrophysiological studies of the agranular cerebellar cortex of ferret after infection with panleukopenia virus have led to the following conclusions: A. At light microscopic level this agranular cerebellar cortex is shown to be totally disorganized when compared to its normal lamination. B. From an ultrastructural point of view, Purkinje cells are present throughout the thickness of the cerebellar cortex, demonstrating dendritic branches studded with unwed spines. C. Purkinje cells receive four different types of inputs-climbing fibers, mossy fibers, stellate cell terminals, and Purkinje cell axon collateral terminals. The mossy fiber contacts which are generally not present in mammals are made in some cases to elongated, “dolichoderic” spines. D. Stellate, basket, and Golgi cell interneurons are present, and receive inputs from mossy and climbing fiber terminals, and probably from axon collaterals of Purkinje cells. The mossy fiber input to these neurons represents a true reorganization of the cerebellar circuit, since it has never been observed in any other vertebrate. E. Stellate and basket cells terminate in contact with dendrites and somata from Purkinje cells, while the Golgi cell terminals seem to be restricted to somata and dendrites of other Golgi cells. F. Electrophysiologically, Purkinje cells can show normal excitability following antidromic invasion, and may be activated by the mossy and climbing fiber afferent systems. Intra-and extracellular recording from Purkinje cells demonstrates that the mossy fiber afferent system activates Purkinje cells with a latency of approximately 1 msec and its excitatory action is graded with relation to the amplitude of the white matter stimulation. Following an initial excitation, this input generates a longlasting inhibition (50 msec), which is attributed to mossy and climbing fiber activation of inhibitory interneurons. G. Climbing fiber activation generates the typical all-or-none burst response in Purkinje cells extracellularly. Intracellularly, large unitary EPSPs characteristic of this form of activation may be recorded with a latency of 1.5 to 3 msec. This large unitary EPSP seems to behave in the usual one-to-one relation (one climbing fiber to one Purkinje cell). H. Double climbing fiber activation reveals that a conditioning white matter stimulation produces a total inhibition of a climbing fiber burst at 15 msec interval, which indicates a rather strong inhibitory action on Purkinje cells following this form of activation. I. Climbing fiber activation is followed in most cases by a so-called climbing fiber reflex, suggesting that the inferior olive in this agranular condition is functioning in a normal manner. J. Finally, some of the neurobiological implications of these findings are considered in the discussion.

The neuronal basis of behavior in Tritonia. IV. The central origin of a fixed action pattern demonstrated in the isolated brain.

Abstract: The mechanisms which initiate and integrate the patterned sequence of activity in the interacting cell groups responsible for the swimming behavior of Tritonia remain operative after complete isolation of the brain from the periphery. Following isolation, a short electrical stimulus applied to a nerve from the oral veil leads to patterned bursting in the flexion neurons of the pedal ganglia, which is identical to that occurring in the same cells during the swimming escape response. Direct evidence of cell-cell interaction in the form of unitary postsynaptic potentials was not obtained, but the close synchronization of burst patterning in both ipsilateral and contralateral synergists infers the existence of positive feed back between many neurons of the group. Alternating bursts of impulses in functional antagonists are associated with reciprocal inhibition between the two networks. As all possibility of proprioceptive feedback has been excluded in the isolated brain, it is evident that the motor output pattern for the escape-swimming response has been generated by a central mechanism without reference to sensory input, which serves only to trigger the action.

Effects of lidocaine on axonal morphology, microtubules, and rapid transport in rabbit vagus nerve in vitro.

Abstract: Treatment with lidocaine, at exposures which completely block impulse conduction and rapid axonal transport, resulted in a sequence of morphological effects on rabbit vagus never in vitro. Low exposures (0.3% for 90 min, 0.4% for 45 min, 0.6% for 25 min) caused a reorientation and proliferation of smooth endoplasmic reticulum (SER); the number of axonal microtubules either remained normal or was increased. Intermediate exposures (0.4% for 90 min, 0.6% for 45 min) Produced a similar effect on the SER, as well as causing a 50% reduction in microtubule number and some swelling of axons and Schwann cells. The highest exposure (0.6% for 75 min) caused over 90% reduction in microtubule number, a partial loss of neurofilaments, and severe swelling of axons, Schwann call, and mitochondria. Reversibility of these effects was tested in lidocaine-treated nerves that had been washed with fresh culture medium. There was good recovery of nerve exposures. After intermediate exposures, there was only partial return of rapid transport, even though imposures, there was morphological, and no recovery occured throughout the excised vagus nerve in efferent and afferent axons as well as Schwann cells, and these events did not coincide with changes in the functional state of rapid axonal transport.

Effects of axonal injury on norepinephrine, tyrosine hydroxylase and monoamine oxidase levels in sympathetic ganglia.

Abstract: A combined morphological and biochemical study was made of the responses of lumbar sympathetic ganglion cells to ligation of their axons in the sciatic nerve of the rat. There was a progressive accumulation of norepinephrine proximal to the ligature for the first few days without detectable changes in the ganglion cells. However, three days after ligation there was a decrease in the axoplasmic accumulation of norepinephrine, accompanied by pronounced falls in ganglion norepinephrine, tyrosine hydroxylase, and monoamine oxidase. These changes, that coincided with the development of signs of chromatolysis in the light and electron microscope, were fully developed one week after ligation and by one month were almost reversed. Proteolysis and death of chromatolytic cells may have made some contribution to the observed changes, but there were no significant changes in the soluble protein content of the lumbar ganglia, presumably because of the counter-balancing increase in protein synthesis known to occur during chromatolysis and regeneration. It is suggested that axotomy caused a reordering of the priorities of protein synthesis in sympathetic neurones to favor the production of proteins necessary for regenerative rather than transmitter functions.

Mechanisms involved in spreading depression.

Abstract: Application of unlabelled glutamate to the isolated chicken retina caused an increase in the tissue transparency and a release of labelled glutamate with which the retina had previously been charged. The threshold for both responses was about 0.2 mM. Stimulation with KCI and direct current elicited transparency changes at a threshold of 8 mM and 4–6 V, respectively, and a release of the label at higher KCl concentrations (75 mM) and potentials (15 V). The transparency change caused by glutamate stimulation was severely depressed by MgC12 (10–15 mM) in contrast to the changes elicited by KC1 and direct current which were little affected by Mg ions. It was postulated that glutamate causes transparency changes by acting on neuronal elements. KC1 would act on glia and would only in high concentrations affect neurons. It was furthermore suggested that the propagation of spreading depression is dependent both on the release of glutamate from the intracellular compartment and on the depolarization of neuronal elements resulting from this release.

Neural mechanisms underlying behavior in the locust Schistocerca gregaria. II. Integrative activity in metathoracic neurons.

Abstract: Preliminary studies are reported on the roles of some interneurons and motorneurons in generating simple behavior patterns in the locust Schistocerca gregaria. The neurons were penetrated by intracellular electrodes singly, or in pairs, whilst a variety of reflexes and spontaneous movements was observed. For example: large oscillations of membrane potential occurring at characteristic flight frequency were observed in flight motorneurons after blowing on the head. The pattern of impulses was directly determined by the amplitude and period of the oscillations. Similar oscillations were observed to occur spontaneously in leg motorneurons, where they caused bursts of impulses which closely resembled those occurring during walking. Both sensory-integrating and pre-motor interneurons were encountered, but none with the function of “commanding” motor output. Much of the input to leg motorneurons is already in programmed form.

Slow accumulation of choline acetyltransferase in crushed sciatic nerves of the rat.

Abstract: Choline Acetyltransferase (CHAc) activity has been estimated using a microradiochemical method in short segments of rat sciatic nerve at different times after crushing the nerve once or with a second crush 2 cm distal to the first. Above a single or proximal crush, the CHAc levels rose slowly in the 5 mm immediately above the crush to reach about 1 1/2 times control at 24 hr. Distal to a single crush and between two crushes no change was detected in CH-Ac-activity up to 18–24 hr. The results are discussed in relation to the rapid accumulation of acetylcholine (ACH) above a crush, and redistribution between two crushes which have previously been demonstrated in the same preparation. It is concluded that the rate of transport of CHAc is probably much slower than that of ACH, and that their mechanismes of transport are not the same.

Electrophysiological analysis of the actions of strychnine, bicuculline and picrotoxin on the axonal membrane.

Abstract: Strychnine, bicuculline, and picrotoxin show similar effects on the membrane of the lobster giant axon. At concentrations above 5 × 10−5 M these compounds cause depolarization of the resting potential and a broadening of the action potential contour. Also occurring is an increase in excitability as defined by a decrease in stimulus threshold. Repetitive firing of action potentials is sometimes observed during the depolarizing process. Drug levels of 5 × 10−4M and higher may secondarily induce depolarizing inactivation of the spike generating mechanism. The loss in resting potential is accounted for by a shift in relative membrane selectivity away from potassium and towards sodium. Associated with this alteration in selectivity is a marked fall in total conductance and abolition of delayed rectification. The results indicate that strychnine, bicuculline, and picrotoxin cause potassium inactivation and further that this effect is manifest in both the resting and active electrogenic state of the membrane.

Neurite development in vitro: II. The role of microfilaments and microtubules in dibutyryl adenosine 3′,5′‐cyclic monophosphate and nerve growth factor stimulated maturation

Abstract: Dibutyryl cyclic AMP (DBC) and Nerve Growth Factor (NGF) have both been shown to enhance in vitro neuronal maturation in embryonic sensory ganglia. Since microfilaments and microtubules are known to be involved in neurite elongation, embryonic chick dorsal root ganglia have been treated with cytochalasin B and colcemid in the presence of these growth promoters (DBC or NGF). Established 48-hr cultures show drastic neurite retraction when treated with either inhibitor. The treatment of cultures with DBC or NGF in the presence of colcemid reverses the inhibitive effects and normal elongation occurs. The growth stimulators, however, are unable to effectively reverse the action of cytochalasin B. These data suggest that increased microtubule assembly is involved in DBC and NGF stimulation of neurite elongation, and hence provide further support for the suggestion that cyclic AMP mediates some of the actions of NGF.

Fast axonal transport in the C-fibers of the garfish olfactory nerve

Abstract: The application of aqueous solutions of tritiated L-Leucine to the olfactory epithelium of the longnosed garfish (Lepisosteus osseus) results in a rapid sorption of the labeled precursor and the subsequent establishment along the nerve of a profile of labeled macromolecular constituents. This profile consists of a sharp wavefront, a peak and a saddle region. At 23°C, the wavefront enters the nerve 1.8 hr after the isotope application and moves away from the cell bodies at an average rate of 222 ± 2.4 mm/day. As the labeled material is propelled along the nerve fiber, its distribution undergoes several characteristic profile changes. The peak broadens and decreases in height and the slope of the wavefront becomes gradually less steep, while the area of the profile increases. These systematic, time-dependent changes suggest a redistribution of some of the labeled axonal material during transport.

The effect of different types of axonal trauma on the synthesis and transport of amine storage granules in rat sciatic nerves.

Abstract: In crushed sciatic nerves of rat the accumulation of noradrenaline (NA), proximal to a ligation, proceeds approximately linearly up to 48 hr after operation, as was earlier reported. After 48 hr the NA in the Proximal portion ceases to increase. The reasons for this arrest in N.A-accumulation have been studied in the present investigation. Rat sciatic nerves were cut, ligated, or crushed at a low level 1–23 days prior to sacrifice. Two hours before death, the nerves were crushed at a level of about 2 cm proximal to the initial operations. Using a histochemical fluorescence method, the relative accumulation of NA above the second high crush was studied. In previously cut or ligated nerves, the NA accumulation observed was very low or zero by 7–9 days, and remained low during the observation period. In previously crushed nerves the NA accumulation was also decreased to rather low levels by 7–9 days. At 15 days, however, the NA accumulation was larger again, and histochemically normal accumulation was observed 17–23 days after the initial crush operation. The results may indicate that axonal trauma causes a depression in the synthesis and transport of amine stronge granules in the adrenergic neurons. Furthermore, the degree and duration of depression seemed to vary with the type of trauma performed. Crushing the nerves, which leaves the connective tissues sheaths uninterrupted, facilitating thus regeneration, appeared to cause a less marked and shorter-lasting depression than cutting or ligating the nerves.

Characterization of fast and slow transported proteins in dorsal root and sciatic nerve of cat.

Abstract: Labeled components, carried by axoplasmic transport into the cat dorsal roots and sciatic nerves following injection of 3H-leucine into the lumbar seventh (L7) dorsal root ganglia, were studied. The pattern of activity present in the subcellular fractions was similar for the two branches of the ganglion neurons entering the nerve and the dorsal root. Fractionation of the proteins present in the high speed supernatant fraction shows that slow transport carries a major proportion of high molecular weight soluble proteins. Further fractination of the soluble proteins with Sephadex G-200 showed labeled activity present in three peaks: Ia, proteins of 480,000 M. W., lb, proteins of 60,000–120,000 M. W. containing colchicine binding (microtubular) protein, and peak II polypeptides, 5,000–10,000 M. W. Isoelectrofocusing of la proteins gave three peaks at pl 4.0, 4.05, and 4.15, and of lb proteins gave peaks at pl 4.0, 4.45, and 4.8 with several minor peaks also present. A gradual shift in pl values of the labeled proteins from acidic to neutral pH was seen from 3 hr to 7 days after the injections. A small amount of soluble proteins and some free leucine is carried down by fast transport, which is characterized by a high amount of activity in the particulate component. A comparison of the distribution of labeled incorporation in the crest and plateau region of fast transported materials showed a somewhat smaller percentage of activity in the small particulate fraction in the plateau, with more labeling of the soluble fraction and with no free leucine present. The results are in conformity with the concept of a compartmentalization of incorporated proteins in the cell body from which materials of somewhat different compositions later enter the axon for transport.

Experimental inhibition of myelination in spinal cord tissue cultures: enzyme assays.

Abstract: Seven specific enzyme activities were assayed in embryonic mouse spinal cord cultures during normal development and under conditions of myelination inhibition and disinhibition. Sulfatase A activity in the inhibited and disinhibited cultures was slightly lower than in the normal control cultures. No specific differences for glucose-6-phosphate dehydrogenase, β-galactosidase, β-glucuronidase, acid prosphatase, and NADP-isocitrate dehydrogenase activities were found among these three experimental conditions. However 2′,3′-cyclic nucleotide phosphohydrolase, an enzyme which has been specifically associated with myelin, showed identical patterns of inhibition and disinhibition as were previously observed for sulfatide synthesis.

Surface culture of fetal mammalian brain cells: Effect of subculture on morphology and choline acetyltransferase activity

Abstract: Cells from brains of fetal rats were grown in surfece cell culture for up to 46 days and were subjected to two subculturing cycles with mild trypsin treatment. The cells maintained their characteristic morphology into third passage, and some putative neurons developed the ability to abhere directly to plastic surfaces. The specific activity of choline acetyl-transferase reached higher levels in third than in first passage cultures. An increase of at least 34-fold was obtained in the total activity of enzyme, the major increments occurring during first and third passages. The data suggest that choline acetyltransferase-producing cells may be capable of multiplication in culture.

Chemosensitivity of mouse neuroblastoma cells in vitro.

Abstract: Mouse neuroblastoma cells have been tested for their responsiveness to iontophoretically applied acetylcholine, norepinephrine, dopamine, and serotonin using intracellularly recorded membrane potential and resis tance as indicators of response Acetylcholine can elicit either a membrane depolarization (D-response) or hyperpolarization (H-response) or a compound D-H response from different neuroblastoma cells of a given culture Some 60 to 70% of tested cells gave no measurable response A change of culture medium (from DMEM to F-12) increased the incidence of H-responses 2-fold Dopamine elicited H-responses in about one-quarter of cells tested, while no responses were elicited by norepinephrine or serotonin in a small sample of cells

Neuronal basis of behavior in Tritonia. II. Relationship of muscular contraction to nerve impulse pattern

Abstract: Recordings were made of the movements elicited in the body of Tritonia diomedia by direct intracellular stimulation of identified brain neurons, which have a motor command function. It was shown that the latency of a movement response decreases as the frequency of stimulation is increased in the range 1 Hz to 20 Hz. Over the same range, the speed of the response increases. The extent, and also the speed, of a movement are proportional to the total number of stimuli, but response speeds and magnitudes vary greatly at different times. The minimum latency, after deduction of conduction time, is 160 ms at 11 °C. It is concluded that the identified brain cells, which cause discrete movements, do so indirectly, by activating a peripheral nerve net which in turn excites the muscle fibers. The nerve net is the site of independent excitatory, inhibitory, and local reflex action.

Dopaminergic neurons in explants of substantia nigra in culture.

Abstract: Explants of substantia nigra and corpus striatum obtained from newborn rats were maintained in tissue culture for up to six days. Explants of substantia nigra exhibited a net increase in the ability to take up H3-dopamine, a process associated with the dopaminergic neurons; in contrast, the explants of corpus striatum showed a rapid loss in this ability to accumulate H3-dopamine. After three days in culture, the specific activity of tyrosine hydroxylase and monoamine oxidase had decreased 50% in explants of substantia nigra. A medium including fetal calf serum and chick embryo extracts was necessary for the increase in H3-dopamine uptake, and nerve growth factor had an inhibitory effect. Histofluorescent examination of nigral explants cultured for three days indicated morphologically normal dopaminergic neurons.

Intracortical impedance changes during spreading depression.

Abstract: Specific impedance at various depths in the cerebral cortex of the rat was measured using a newly developed thin film coaxial microelectrode. This novel electrode has the advantage of measuring specific impedance of extremely small volumes of tissue, and thus of making highly localized measurements. The resting level of cortical impedance in the rat was found to be stable, but to vary for different sites and animals from 200–700 ohm-cm. During spreading depression, loop-shaped impedance loci were consistently observed in the complex impedance plane with peak magnitude 2–3 times resting level. These unusual loop-shaped loci are thought to represent distinct process differences during development and recovery phases of spreading depression. This preliminary investigation has demonstrated the feasibility of a technique for making highly localized impedance measurements in nervous tissue. Such measurements are useful for in vivo studies of such tissue parameters as extracellular space.

Restorative effects of cyclic AMP on complex bioelectric activities of cultured fetal rodent CNS tissues after acute CA++ deprivation.

Abstract: Explants of fetal mouse spinal cord and cerebral cortex generate organotypic slow-wave and repetitive-spike discharges in vitro which can be abolished by agents which reduce the concentration of Ca++ available to the tissue. Synaptically mediated discharges are rapidly blocked in Ca++-free balanced salt solution (BSS), or in regular BSS after addition of 10−3 M EGTA, 5–10 × 10−3 M Mg++, or 10−4 M xylocaine, but simple spike potentials can still propagate. When low concentrations of cyclic AMP or dibutyryl cyclic AMP (2 × 10−6 M) are added to the Ca ++-free BSS or Ca++-antagonist-BSS, a temporary (1–20 min) restoration of characteristic complex bioelectric activities occurs (or the onset of depression is delayed if cyclic AMP is initially added). Phosphodiesterase inhibitors, e.g. 10−3 M caffeine, are also effective in restoring these blockades, whereas 5′AMP and ATP are not. Application of 10−6 M cyclic AMP or 10−3 M caffeine in regular BSS greatly enhances excitability of some CNS explants, resembling convulsive effects observed in CNS in situ. The data suggest that cyclic AMP can mobilize Ca++ from membranebound Ca pools within neurons in CNS explants so as to permit Ca++-dependent release of neurotransmitter during Ca++ deficits. Thus, it may also be that under normal conditions, cyclic AMP can regulate the availability of Ca++ for synaptic transmission in the central nervous system, thereby modulating the efficacy of synaptic functions.

Genetic manipulation of motor output in shaker mutants of Drosophila

Abstract: Hk1, Hk2, Sh5, and Eag are sex-linked behavior mutants of Drosophila which shake their legs vigorously when etherized The shaking rate increases with age to a plateau by five days after eclosion; it is decreased by increasing ether concentrations Eag is temperature sensitive, shaking vigorously at 30°C but not at all below 20°C Hk1 and Hk2 have cyclic shaking patterns consisting of three to six periods of shaking per minute Hk1 stakes more, and has longer shaking periods than Hk2; the heterozygote Hk1/Hk2 is intermediate When heterozygous with a deletion for the locus, both Hk1 and Hk2 shake more, the pattern of Hk2 resembling that of Hk1 Therefore, these alleles may differ only quantitatively The shaking pattern of Sh5 is distinctly different, consisting of short bursts of shaking two or three times a second Flies with both Hk and Sh5 mutations in different combinations can have a pattern which is Hk-type, Sh5-type, or both simultaneously, depending upon the geno-type The shaking of Hk1 is known to be a consequence of abnormally firing neurons in the thoracic ganglion Therefore, the interactions between Hk and Sh5 are probably between two different mutant effects upon the motor system, both subject to genetic manipulation

Axoplasmic transport: differential inhibition by cytochalasin-B.

Abstract: Slow (1 mm/day) and fast (10mm/day) axoplasmic transport of labelled protein, shown by the movement of sharp peaks of radioactivity along the crayfish nerve cord, are blocked by cytochalasin-B. Labelled proteins in the slow peak maintain their sharp distribution; however, those in the fast peak disperse over a broad area after the drug-induced blockage.

Interconversion of glycine and serine in a synaptosome fraction isolated from the spinal cord, medulla oblongata, telencephalon, and cerebellum of the rat.

Abstract: The metabolism of glycine was studied in vitro using preparations of synaptosomes (P2) isolated from the spinal cord, medulla oblongata, telencephalon, and cerebellum of the rat The content of aspartate, glutamate, glutamine, serine, threonine, glycine, and alanine was determined in the crude synaptosomal fraction (P2) from the four regions of the CNS In general, the content of the amino acids in the subcellular fraction from the medulla oblongata and spinal cord resembled one another whereas the same was true for the telencephalon and cerebellum Significantly greater amounts of glutamate, aspartate, glutamine, and serine were found in the synaptosomal fraction of the telencephalon and cerebellum than in similar P2 fractions from the medulla oblongata and spinal cord On the other hand, glycine had a higher content in the latter two preparations than in the P2 fractions from the telencephalon and cerebellum [U-14C]-serine and [U-14C]-glycine were readily taken up by the crude synaptosomal fractions The rate of conversion of serine to glycine appeared to be 185, 39, 29, and 27 times greater than the conversion of glycine to serine in the incubated synaptosomal fractions from the cerebellum, spinal cord, medulla oblongata, and telencephalon, respectively The conversion of serine carbons into glycine appeared to be 25, 19 and 16 times greater in the synaptosomal fraction from the cerebellum, spinal cord, and medulla oblongata than in similar preparations from the telencephalon The data are discussed in terms of the possible synthesis pathway for the pool of glycine used as a transmitter

Axoplasmic transport of RNA.

Abstract: The transport of RNA from the ganglion cell bodies within the retina to the contralateral optic tectum has been studied in the chick following intraocular injection of radioactive uridine. By tracing the appearance of labeled RNA at the proximal end of the optic nerve as it leaves the eyeball and comparing this to the time of arrival of RNA within the optic tectum, the migratory velocity of axonal RNA has been calculated to be around 12 mm per day. The continuation of RNA migration to the optic tectum in the presence of intracerebrally injected actinomycin-D but not in the presence of the intraocularly injected drug, suggests a retinal site of synthesis of the excess RNA found in the tectum innervated by the injected eye. A study of the rate of disppearance of radioactivity of the transported RNA in the optic lobes, suggested that this RNA turns over more rapidly than the bulk of tectal RNA. The destination of migrating RNA within the optic tectum has been autoradiographically examined. Most radioactive RNA is found in the outer tectal layers in which are found the afferent fibers of the optic tract and most of their synaptic terminations. Label is not confined to these areas however but is also present in the deeper layers of the optic tectum which are not known to contain any primary synapses of the axons from retinal ganglion cells.

Transport of sulfate in the rabbit's brain

Abstract: The possibility that, in the rabbit, sulfate is actively transported across the choroid plexuses was investigated by ventriculo-cisternal perfusion in the absence and presence of carrier. No evidence for this could be found. Penetration of sulfate from blood to brain and CSF was measured in the intact animal equilibration was very slow, indicating the presence of high barriers for this solute, but the low steady-state level in the brain, approached in these experiments, was quite consistent with slow passive permeability aacross the barriers, with free exchange between extracellular fluid and CSF. The clearance of another slowly penetrating solute, creatinine, from the perfusion fluid during ventriculo-cisternal perfusion was comparabl with that of sulfate.