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F Strumwasser

Publications and source records attributed to F Strumwasser.

At least 37 records · Page 2Linked to original sources
PubMed1980

Purification and primary structure of two neuroactive peptides that cause bag cell afterdischarge and egg-laying in Aplysia.

Two neuroactive peptides, A and B, have been isolated from the atrial gland in the reproductive tract of Aplysia. Each of the two peptides is able to induce egg-laying behavior in recipient animals. In vitro recordings from the abdominal ganglion show that both peptides also trigger longlasting discharges in the bag cell neurons at concentrations around 0.1 muM. The peptides were purified by a combination of ammonium sulfate precipitation, agarose gel filtration, and cation exchange chromatography. Each peptide has 34 amino acid residues. Microsequencing together with carboxypeptidase Y degradation and analysis of tryptic peptides revealed the following sequence for peptide A: H-Ala-Val-Lys-Leu-Ser-Ser-Asp-Gly-Asn-Tyr-Pro-Phe-Asp-Leu-Ser-Lys-Glu-Asp-Gly -Ala-Gln-Pro-Tyr-Phe-Met-Thr-Pro-Arg-Leu-Arg-Phe-Tyr-Pro-Ile. Peptide B differs from A in only four positions. The first nine residues of B are: Ala-Val-Lys-Ser-Ser-Ser-Tyr-Glu-Lys-, whereas residues 10-34 of B are identical to those of A. The calculated M(r) of A is 3924 and that of B is 4032. The pI of peptide A as determined by isoelectric focusing in polyacrylamide gels is 7.9-8.1 and that of peptide B is 9.0-9.2. It is estimated that each atrial gland contains at least 150 mug of peptide A and 50 mug of B. Neither peptide resembles the egg-laying hormone isolated from bag cell neurons. It is postulated that the atrial gland peptides are released during copulation, and then by interacting with neuronal receptors in the head ganglia and pleuroabdominal connectives they cause the bag cells to afterdischarge, thereby releasing egg-laying hormone.

Action Potentials↗
PubMed1980

The differential effects of ionizing radiation on the circadian oscillator and other functions in the eye of Aplysia.

Ionizing radiation has been used to selectively separate the circadian oscillator function of the eye of Aplysia from some of its other functions--synchronous compound action potential (CAP) generation, the light response, synaptic transmission between photoreceptors and output neurons, and the bursting pacemaker mechanism. Doses of 4-krad (50 kV peak) x-rays have a minimal effect on the circadian rhythm of CAP frequency, measured from the otpic nerve, whereas irradiation with a 40-krad dose abolishes the rhythm without affecting any of the four other functions of this eye (1 rad = 0.01 J/kg = 0.01/Gy). We estimate a 50% survival of the oscillator function at doses of about 6 krad. The oscillators of irradiated eyes are not merely desynchronized when the rhythm is abolished, because in vitro light-dark entrainment does not restore free-running rhythmicity. The results, including those from selective irradiation of the anterior or posterior poles of the eye, suggest that there are a number of circadian oscillators in the eye--most of them in the posterior portion near the optic nerve. An approximate target size has been obtained from target theory approximately equal to 10(8) A3, which is somewhat larger than the target size for viral infectivity function, as one example. There are reservations about estimating target size in a complex organ such as the eye. However, this approximate target size and the fact that recovery or repair can occur in vivo suggest that the oscillator may involve nucleic acid molecules.

Action Potentials↗
PubMed1980

Neurosecretion of egg-laying hormone and other peptides from electrically active bag cell neurons of Aplysia.

1. Radiolabeled peptides released from an isolated cluster of bag cell neurons, during an after discharge, were compared with the polypeptide egg-laying hormone (ELH), 4,500 daltons, pI 9.0-9.3, as purified from homogenates of bag cell clusters. A peptide, labeled with methionine, leucine, and arginine, which is selectively released from after discharging bag cell clusters comigrates with marker ELH, purified from cluster homogenates, on P-6 gel filtration columns and subsequent isoelectric focusing gels. At least three other presumed peptides of unknown function are also released, including one of 5,000-6,000 mol wt, pI 4.5-5.0. 2. When bag cells afterdischarge in vitro, bioactive material is released that will induce egg laying when injected into an Aplysia. This released bioactive material also comigrates with bioactive material from cluster homogenates on P-6 columns. 3. These experiments demonstrate that ELH (4,500 mol wt, pI 9.0-9.3), as purified from bag cell cluster homogenates, is also the major form secreted from bag cells that induces egg laying. This purified ELH can now be used to study the physiological effects of a secreted neurohormone and their relationship to behavior.

Action Potentials↗
PubMed1980

Neuronal sites of action of a neurosecretory peptide, egg-laying hormone, in Aplysia californica.

1. Egg-laying hormone (ELH) is a polypeptide of about 4,500 mol wt synthesized in the bag cell neurons of the abdominal ganglion of Aplysia. We studied the effects of ELH on the neuronal activity of the attached head ganglia (buccal, cerebral, pleural, and pedal), on the isolated buccal ganglia, as well as on feeding in intact Aplysia. 2. Starved animals (n = 7) injected with crude extract containing ELH stopped eating algae at 17 +/- 4 min and their eggs first appeared at 29 +/- 4 min after injection at 20 degrees C. This cessation of eating is significant when compared to the seven controls (P less than 0.01). These data clearly indicate that a suppression of feeding activity occurs before the appearance of eggs. 3. ELH applied to the paired buccal ganglia in vitro activates a pair of neurons into a tonic pacemaker mode (approximately 1 spike/s). This activation also occurs in a high-magnesium, zero calcium solution that blocks chemical synapses. The time for the full appearance of this activity in vitro correlates well with the time for suppression of feeding in vivo. Each of these neurons has an ipsilateral axon in buccal nerve 3. The neuron has been identified by intracellular recording. 4. ELH increases the rate of firing of a second pair of buccal neurons, each with an ipsilateral axon in the cerebrobuccal connective. 5. ELH, when applied to the attached head ganglia, causes large bursts of neuronal activity in pedal nerves to the foot and increased activity in the nerve to the penis; the relevant neurons remain to be identified. 6. These in vitro effects were produced by ELH partially purified from bag cell cluster homogenates using ammonium sulfate precipitation followed by anion exchange and gel filtration chromatography or by ELH released from activated bag cells in isolated abdominal ganglia and then purified by gel filtration. The isolated buccal ganglia effects have been confirmed with fully purified ELH. 7. The ELH effects on the in vitro nervous system support the hypothesis that ELH in vivo acts directly on the nervous system to suppress feeding activity, controlled by the buccal and cerebral ganglia. ELH may also produce characteristic movements of the head during egg laying, controlled probably by the pedal and cerebral ganglia.

Animals↗
PubMed1980

Peptides controlling behavior in Aplysia.

Figure 11 summarizes our present understanding of the relationships between the bag cells, the atrial gland, their respective peptides, the central nervous system, and reproductive behavior. There are some interesting aspects of the overall organization of the system. The three hormonal peptides (ELH and the two atrial gland peptides) have specific actions on the central nervous system not unlike what we are currently learning from mammalian systems (e.g., LHRH and TRH). ELH, in addition, has several specific peripheral targets, the details of which remain to be worked out. The fact that ELH and other hormones have multiple targets within the central nervous system as well as nonnervous peripheral targets raises the question of whether one or more different receptors exist for single hormone. We suggest that peptides larger than perhaps five residues may carry several "messages" or receptor binding sites encoded within the one molecule. Large peptides such as ELH could obviously have separate domains of charge distribution within the molecule, and these would have the advantage, over the classical small molecule transmitters, of activating a variety of very different targets. The atrial gland is a peripheral source of peptides with potent nervous system actions; this is reminiscent of peptides in mammals, e.g., substance P, gastrin, and somatostatin, all of which were initially isolated from the gut and which are now being found in and also have actions on the central nervous system. Such resemblances in the principles of organization between mammals and molluscs are constant reminders that neuropeptidergic systems are old tricks in the evolutionary bag and that what we learn from molluscs and other invertebrates about mechanisms and organization will likely apply to mammals.

Amino Acid Sequence↗
PubMed1979

The morphology and coupling of Aplysia bag cells within the abdominal ganglion and in cell culture.

The bag cells in the abdominal ganglion of Aplysia californica control egg-laying behavior by releasing a polypeptide (ELH) during an afterdischarge of synchronous action potentials. We have used intracellular injection of Lucifer Yellow to study the morphology and interconnections of the bag cells. These neurosecretory cells are typically multipolar and their processes extend in all directions out from the bag cell clusters into the surrounding connective tissue, where they branch in a complex manner. In some of the dye injection experiments, dye transfer from the injected cell to neighboring cells was observed. Freeze fracture of the bag cell clusters and their surrounding connective tissue revealed numerous gap junctions on bag cell processes within the clusters as well as on more distal processes. We have also examined the morphology and coupling between bag cells in primary culture. As in the intact ganglion, bag cells in culture were found to be multipolar. All pairs of bag cells whose somata or processes had formed contacts in culture were electrically coupled. The strongest coupling was observed between pairs of cells whose somata appeared closely apposed. In these cases transfer of Lucifer Yellow between cells could also be observed. It is therefore likely that the synchrony of bag cell action potentials during a bag cell afterdischarge is a result of coupling between individual cells in the bag cell cluster.

Animals↗
PubMed1979

Purification and primary structure of the neuropeptide egg-laying hormone of Aplysia californica.

Egg-laying hormone (ELH), a neuropeptide synthesized by the bag cell neurons, induces egg laying and its correlated behavior in Aplysia californica. In the present study, ELH has been purified to homogeneity and its primary structure has been determined. We find this molecule to have 36 amino acid residues with a M(r) of 4385 and a calculated isoelectric point of 9.7. Direct microsequence analysis revealed a single amino acid sequence that is in agreement with the amino acid composition determined after acid hydrolysis of ELH: H-Ile-Ser-Ile-Asn-Gln-Asp-Leu-Lys-Ala-Ile-Thr-Asp-Met-Leu-Leu-Thr-Glu-Gln- Ile-Arg-Glu-Arg-Gln-Arg-Tyr-Leu-Ala-Asp-Leu-Arg-Gln-Arg-Leu-Leu-Glu-Lys-OH. Enzyme data indicate that the COOH-terminal lysine may be modified but its exact nature remains to be determined. There is no similarity between the amino acid sequence of ELH and that of presently known vertebrate neuropeptides. The two-step purification procedure, starting with a homogenate of bag cell clusters, consisted of cation exchange chromatography on SP C25 (Sephadex) followed by gel filtration on Bio-Gel P-6. Our purification results in a 100-fold enrichment of ELH from bag cell homogenates and a 36% recovery of purified radiolabeled marker ELH. Analysis of purified ELH radiolabeled with [(35)S]methionine or [(3)H]leucine on isoelectric focusing gels and on 8 M urea/sodium dodecyl sulfate gels showed only a single peak containing 90% of the radiolabel. Radiolabeled ELH migrated with a pI of 9.0-9.2 and an apparent M(r) of 3500-5700. ELH retained egg-laying bioactivity when eluted from this segment of the gel. We find that 2.5 nmol of pure ELH consistently induces egg laying at 20 degrees C.

Amino Acid Sequence↗
PubMed1978

Membrane-potential-sensitive dyes for optical monitoring of activity in Aplysia neurons.

Two membrane-associated dyes (WW375 and NK2367) which change their absorption of light when the membrane potential changes have been studied using several preparations form Aplysia. Action potentials are easily observed in nerve trunks (from a number of axons), in bag cell clusters, in some of the larger single cells of the parietovisceral ganglion, and in the optic nerve. Physiological effects of the dyes on the circadian rhythm of activity in the eye are described.

Action Potentials↗
PubMed1978

Neurotransmitter modulation, phosphodiesterase inhibitor effects, and cyclic AMP correlates of afterdischarge in peptidergic neurites.

The neuroendocrine bag cells in the abdominal ganglion of Aplysia generate a long-lasting synchronous afterdischarge upon brief stimulation of an afferent pathway. After this afterdischarge the cells become refractory to further synaptic stimulation. We find that synchrony, afterdischarge, and prolonged refractoriness are properties that can be expressed in the isolated asomatic neurites of the bag cells. We have distinguished two independent types of refractoriness. The first (type I) is seen as a failure of action potentials generated in the tips of bag cell neurites to invade cell somata. The second form of refractoriness (type II) controls the duration of afterdischarge such that stimuli after the first afterdischarge produce only very short afterdischarges or fail to elicit an afterdischarge. Type II refractoriness is sensitive to serotonin and certain of its analogues, and to dopamine and the methylxanthine phosphodiesterase inhibitors. Extracellularly applied serotonin suppresses an ongoing afterdischarge while dopamine and the phosphodiesterase inhibitors, when applied at the end of the first afterdischarge, generate a subsequent afterdischarge of long duration without further electrical stimulation. None of these compounds influenced the degree of type I refractoriness. We have shown that both serotonin and dopamine stimulate the formation of cyclic AMP in the bag cell clusters and in the pleurovisceral connectives and that the occurrence of an afterdischarge is associated with a specific increase in total cyclic AMP in bag cell bodies. Moreover, afterdischarges can be generated in unstimulated preparations by extracellular application of the cyclic AMP analogues, 8-benzylthio-cyclic AMP or 8-methylthio-cyclic AMP. Our data suggest that serotonin and/or dopamine may control bag cell activity and that activation of adenylate cyclase is linked to bag cell afterdischarge.

Action Potentials↗
PubMed1976

Patterns of proteins synthesized in the R15 neuron of Aplysia. Temporal studies and evidence for processing.

The time-course of changes in the pattern of newly synthesized proteins in the R15 neuron of the parietovisceral ganglion of Aplysia californica has been studied at 14 degrees C. 5% polyacrylamide gels containing sodium dodecyl sulfate (SDS) have been used to separate newly synthesized (leucine-labeled) proteins from the neuron. We have demonstrated that the pattern of newly synthesized proteins from the R15 neuron does not change significantly if 5-h pulses of labeled leucine are given during the first 72 h of in vitro incubation of the excised ganglion. However, the level of leucine incorporation begins to decline somewhere between 17 and 43 h after the ganglion is isolated; at 43 and 69 h the levels of incorporation fell to 29 and 10% of the initial level, respectively. A number of conclusions have been drawn from the use of a sequential, double-label type of experiment in the same cell. There is processing of SDS-soluble, 12,000-dalton (12k) material to 6,000-9,000-dalton (6-9k) material. These materials are the two major peaks on gels after long labeling periods and together account for about 35% of all newly synthesized proteins. After synthesis of 12k material, there is a gradual disappearance of 12k (half-life about 8 h) and simultaneous appearance of 6-9k material on the gels, as the postsynthesis "chase" period of ganglia incubation is increased. The processing of 12k to 6-9k material occurs even in the presence of anisomycin, a protein syntehsis inhibitor, during the chase period. While the rate of 12k to 6-9k conversion can vary from cell to cell, it appears to remain consistent within, and is characteristic of, any individual R15. We detect no circadian rhythm in either the rate of 12k synthesis or the rate of 12k to 6-9k processing with 5-h label periods. These results are discussed in relation to the roles of 12k and 6-9k material in the R15 neuron.

Animals↗
PubMed1976

Phase shifting the circadian rhythm of neuronal activity in the isolated Aplysia eye with puromycin and cycloheximide. Electrophysiological and biochemical studies.

The effects of pulse application of puromycin (PURO) or cycloheximide (CHX) were tested on the circadian rhythm (CR) of spontaneous compound action potential (CAP) activity in the isolated Aplysia eye. CAP activity was recorded from the optic nerve in constant darkness at 15degreesC. PURO pulses (6, 12 h; 12--134 mug/ml) and CHX pulses (12 h, 500--2,000 mug/ml) caused dose-dependent phase delays in the CR when administered during projected night. PURO pulses (6 h, 125 mug/ml) caused phase advances when given during projected day and caused phase delays when given during projected night. In biochemical experiments PURO (12 h, 20 mug/ml) and CHX (12 h, 500 mug/ml) inhibited leucine incorporation into the eye by about 50%. PURO (12 h; 50, 125 mug/ml) also changed the molecular weight distribution of proteins synthesized by the eye during the pulse. The effect of PURO (12 h, 125 mug/ml) on the level of incorporation was almost completely reversible within the next 12 h but the phase-shifted eye showed an latered spectrum of proteins for up to 28 h after the pulse. In electrophysiological experiments spontaneous CAP activity and responses to light were measured before, during, and after drug treatments. In all, eight parameters in three periods were analyzed quantitatively. Of these 24 indices, only 3 showed significant changes. PURO increased spontaneous CAP frequency by 67% 0-7 h after the drug pulse and increased the CAP amplitude of the tonic light response by 23% greater than 7 h after the pulse. CHX increased the intraburst spontaneous CAP frequency by 33% during the pulse and CAP frequency of the tonic light response by 32% 0-7 h after the pulse. The above data indicate that phase-shifting doses of PURO and CHX inhibit protein synthesis in the eye without causing adverse electrophysiological effects, and suggest that protein synthesis is involved in the production of the CR of the isolated Aplysia eye.

Action Potentials↗
PubMed1975

Circadian Rhythm of Neuron R15 of Aplysia californica: In Vivo Photoentrainment.

(1) The neuron R15 in the parietovisceral ganglion of Aplysia has a circadian rhythm of spiking activity when recorded in the isolated ganglion. The rhythm is entrained in vivo by light-dark cycles. (2) The phase of the R15 rhythm is a function not only of the entraining light schedule, but also of the time of dissection. Changes in the dissection time during the light portion of the light-dark cycle yield little change in the subsequent R15 peak time. Dissections during the dark portion produce peak times that vary with dissection time with a slope that is approximately one. (3) The circadian rhythm of R15 can be phase-shifted in vivo by changes in the phase of the entraining light-dark cycle in one to two weeks. R15 neurons of blinded Aplysia, however, show little or no phase shift in this time. (4) It is concluded that the eyes are important as receptors for the photoentrainment of the R15 rhythm in vivo, but that neural connections from the eyes to R15 are not required.

Journal Article↗