Test of the negative feedback hypothesis of colony size sensing in social insects.

Social insects can sense colony size-even without visual information in a dark environment. How they achieve this is yet largely unknown. We empirically tested a hypothesis on the proximate mechanism using ant colonies. In Diacamma colonies, the monogynous queen is known to increase the effort devoted to queen pheromone transmission behaviour (patrolling) as the colony grows, as if she perceives colony size. The negative feedback hypothesis assumes that, through repeated physical contact with workers, the queen monitors the physiological state (fertility) of workers and increases her patrolling effort when she encounters more fertile workers. Supporting this hypothesis, we found that the queen increased her patrolling effort in response to a higher ratio of fertile workers under the experimental condition of constant colony size. Furthermore, chemical analyses and bioassays suggested that cuticular hydrocarbons have queen pheromone activity and can mediate the observed queen-worker communication of fertility state. Such a self-organizing mechanism of sensing colony size may also operate in other social insects living in small colonies.

Speed of back-swimming of Lymnaea.

The pond snail, Lymnaea stagnalis, can locomote on its back utilizing the surface tension of the water. We have called this form of movement 'back-swimming'. In order to perform this behavior, the snail must flip itself over on its back so that its foot is visible from above. Little is known about the mechanism of this back-swimming. As a first step for the elucidation of this mechanism, we measured the speed of back-swimming of Lymnaea at the different times of the day. They back-swam significantly faster in the morning than just before dark. These data are consistent with our earlier findings on circadian-timed activity pattern in Lymnaea. Lymnaea appear to secrete a thin membrane-like substance from their foot that may allow them to back-swim. To confirm the existence of this substance and to examine whether this substance is hydrophobic or hydrophilic, we applied a detergent onto the foot during back-swimming. A single drop of 1% Tween 20 drifted Lymnaea away that were still kept at the water surface. These results suggest that Lymnaea secrete a hydrophobic substance from their foot that floats to the water surface allowing Lymnaea to back-swim.

Promotion of sprouting and synaptogenesis of cerebrofugal fibers by ganglioside application in the red nucleus.

The effect of gangliosides on sprouting of cerebrorubral synapses from the contralateral cerebral cortex was investigated following lesions of normal ipsilateral cerebrorubral synapses. It was found that, in cats in which ipsilateral cerebral ablations were performed at more than 2 months of age, the mean probability of occurrence of EPSPs induced in red nucleus neurons by stimulation of fibers from the contralateral cerebral cortex in the ganglioside-treated group was significantly higher than in the untreated group. Therefore, ganglioside application appears to promote sprouting and formation of functional synapses in the red nucleus.

The possible role of collateral sprouting in the functional restitution of corticospinal connections after spinal hemisection.

We investigated in monkeys whether the corticospinal fibers increase their connections with lumbosacral neurons after spinal hemisection, using the retrograde horseradish peroxidase (HRP) method. In three monkeys 3.5-38 months after spinal hemisection at the lower thoracic or upper lumbar cord, HRP was injected into the lumbosacral cord unilaterally on the hemisected side at a level caudal to the lesion. Control injections were performed in two intact monkeys and in two other monkeys immediately after hemisection. In all animals, corticospinal neurons in the precentral motor cortex were labeled bilaterally. However, in the chronically spinal hemisected monkeys, the number of the labeled neurons was significantly increased on the side ipsilateral to the lesion. These results suggest that corticospinal connections to lumbosacral motoneurons are newly formed on the side of spinal cord hemisection. This synapse formation may be due to collateral sprouting of intact corticospinal fibers, and it may underlie the mechanisms of motor recovery.

Histochemical study on the relation between NO-generative neurons and central circuitry for feeding in the pond snail, Lymnaea stagnalis.

To examine whether nitric oxide (NO)-generative neurons are included in the central circuitry for generation of feeding pattern in the pond snail, Lymnaea stagnalis, two staining techniques for NADPH diaphorase and serotonin (5-HT) were applied for its central nervous system (CNS). The former technique is known to show localization of NO synthase; the latter is well employed as a marker for the feeding circuitry because 5-HT is a main transmitter in it. In the buccal ganglion, B2 motoneuron was found to be a putative NO-generative neuron. This motoneuron is not involved directly in the coordination of feeding pattern but is activated simultaneously with the feeding to control the oesophageal and gut tissues for the digestion. Taking account of the diffusion effects of NO, the NO released from B2 motoneuron, when the feeding is started, is thought to sufficiently modulate the feeding circuitry. In the cerebral ganglion, the superior lip nerve, the median lip nerve and the tentacle nerve included both putative NO-generative fibers and serotonergic fibers. These fibers are not identical, but the NO released in the nerves may activate the serotonergic fibers, resulting in the influence upon the initiation of the feeding. Therefore, our present findings clearly showed that NO is not involved in transmission within the central circuitry for the feeding, but suggested that NO can crucially affect the feeding behavior, such as initiation and modulation of the feeding pattern.

Histological characterization of lip and tentacle nerves in Lymnaea stagnalis.

The lip and tentacle nerves of the pond snail, Lymnaea stagnalis, were characterized using histological techniques. Anatomical drawings showed the detailed distributions of the superior lip, median lip, and tentacle nerves in the lip and tentacle; in particular it was found that the mouth is mainly innervated by the superior lip nerve. The tentacle nerve was clarified to form a zigzag structure along the extension direction in a shrinking tentacle. By backfilling of the superior lip nerve and/or the median lip nerve with fluorescent dyes, the neurons in the CNS made some clusters, whereas those stained from the tentacle nerve made other clusters. These stained neurons were not part of the central pattern generator or its regulatory neurons for feeding. The present results, therefore, suggest that the superior lip nerve may be employed as a principal factor in the chemosensory transduction from the mouth, and that no direct inputs occur through the lip and tentacle nerves to the central pattern generator or its regulatory neurons for feeding.

Physiological characterization of lip and tentacle nerves in Lymnaea stagnalis.

The lip and tentacle nerves of the pond snail, Lymnaea stagnalis, were characterized using electrophysiological techniques. When the activity of those nerves was induced in lip-tentacle preparations, aversive taste signals were transmitted through all the lip and tentacle nerves, but appetitive signals could be recorded only through the superior lip nerve. In the CNS immersed in high Mg2+ -high Ca2+ saline, electrical stimuli applied to any of the nerves failed to induce action potentials in one of the regulatory neurons (cerebral giant cell: CGC) involved in feeding responses, implying that the signals are polysynaptically transmitted to the CGC. Intracellular recordings revealed that the CGCs in semi-intact half-body preparations received both appetitive and aversive taste signals not only through the superior lip nerve but also through the median lip nerve. In addition, an osphradium was ruled out as a candidate for appetitive reception. The present results, together with our preceding data arrived at by the histochemical analyses, indicate that the appetitive taste transduction responsible for generating feeding responses is performed through the superior lip nerve with some contribution of the median lip nerve. The data showing that the CGC can receive various taste signals suggests that it may play a crucial role in feeding behavior as demonstrated in the study of conditioned taste-aversion.

Motor recovery after cross-reinnervation of a forelimb nerve by the phrenic nerve in cats.

Electromyographic (EMG) activities of forelimb muscles and the diaphragm following cross-reinnervation of the elbow flexor nerve by the phrenic nerve sampled simultaneously with cinematographic recording of forelimb movements, in order to examine whether the respiratory nervous system in adult cats is capable of compensation for movement disorders. The right phrenic and musculocutaneous nerves were cut preparatory to cross-union, the proximal stump of the phrenic nerve being joined to the peripheral stump of the musculocutaneous nerve. Reinnervation of the elbow flexor muscles (m. biceps brachii and m. brachialis) was confirmed in terminal experiments by observing muscular contractions induced by nerve stimulations and/or retrogradely labeled motoneurons in cervical (C5-C7) segments following treatment for horseradish peroxidase (HRP) which had been injected previously into the m. biceps brachii of both sides. Behavioral observations were made in the chronically cross-reinnervated cats. Bipolar EMG recording electrodes were implanted chronically in the right m. biceps brachii, m. triceps brachii and in the left intact diaphragm. In freely moving cats 3-22 months after cross-union surgery, EMG activities in the elbow flexor usually were synchronized with those of the left diaphragm. However, occasionally, in cats cross-reinnervated more than 6 months previously, EMG activities in the right biceps muscle were dissociated from those of the left diaphragm during voluntary movements. During walking, EMG activities in the cross-reinnervated biceps muscle were not coordinated with the walking cycle. By contrast, during voluntary goal directed movements, the cats were capable of reaching their right forepaws to the target in a smooth manner using elbow flexion that was associated with EMG activity of the biceps muscles.(ABSTRACT TRUNCATED AT 250 WORDS)

Differential effects of a benzodiazepine on synaptic transmissions in rat hippocampal neurons in vitro.

The effects of midazolam, one of the most popular benzodiazepines, on synaptic transmissions were compared with intracellular recordings between CA1 pyramidal cells (CA1-PCs) and dentate gyrus granule cells (DG-GCs) in rat hippocampal slices. First, we studied the effects of midazolam on orthodromically evoked spikes, membrane properties and synaptic potentials. Secondly, the effects of a GABA(A) receptor agonist, muscimol, were examined on membrane properties to determine whether or not the densities of GABA(A) receptors are different between CA1-PCs and DG-GCs. Midazolam (75 microM) markedly depressed orthodromically evoked spikes in CA1-PCs, compared with those in DG-GCs. A GABA(A) receptor antagonist, bicuculline (10 microM), almost completely antagonized the depressant effects of midazolam on spike generation in CA1-PCs, whereas it had little effect on midazolam in dentate gyrus granule cells. Midazolam produced either depolarizing or hyperpolarizing effects on resting membrane potentials (Vm) with an input resistance decrease in CA1-PCs, whereas it produced depolarized Vm in DG-GCs. Midazolam significantly increased the amplitude of monosynaptic inhibitory postsynaptic potentials in CA1-PCs, whereas midazolam slightly decreased these in DG-GCs. Midazolam significantly decreased the amplitude of excitatory postsynaptic potentials both in CA1-PCs and DG-GCs. Muscimol (100 microM) produced either depolarizing or hyperpolarizing effects on Vm with an input resistance decrease in CA1-PCs, and it depolarized Vm with an input resistance decrease in DG-GCs. These results demonstrate that midazolam has differential effects on excitatory and inhibitory synaptic transmissions in hippocampal neurons. The mechanism of this difference could be partly due to the different types of GABA(A) receptors between CA1-PCs and DG-GCs.

Morphological and electrophysiological study of sprouting of corticorubral fibers after lesions of the contralateral cerebrum in kitten.

The appearance of crossed corticorubral projections following ablations of the ipsilateral cortex is shown to result from the formation of new connections and is not due to the preservation of pre-existing bilateral connections. At least some of these crossed projections are collaterals of the pyramidal tract. Post-tetanic potentiation can be demonstrated both intra- and extracellularly following ipsilateral cerebral peduncle stimulation whereas no changes in excitatory postsynaptic potential amplitude are observed following contralateral cerebral peduncle stimulation.

Complement receptor 3-like immunoreactivity in the light green cells and the canopy cells of the pond snail, Lymnaea stagnalis.

We observed CR3-like immunoreactivity in the central nervous system (CNS) and its surrounding peripheral nerves of the pond snail, Lymnaea stagnalis. In the CNS of L. stagnalis, the immunoreactivity presenting meshwork-like structure was detected in some neurosecretory cells, which are the light green cells (LGCs) and the canopy cells (CCs), both controlling the body growth. The immunoreactivity was also observed along the edges of median lip nerves. The immunoreactive regions in the median lip nerves appeared to form the axonal plates, from which the LGCs and the CCs release molluscan insulin-related peptides (MIPs) into the blood. By contrast, no immunoreactivity was detected in other neurosecretory cells or their release sites, for example the caudodorsal cells and the cerebral commissure, which release ovulation hormones. The present findings, therefore, suggested that CR3 expresses only in the neurosecretory cells releasing MIPs in L. stagnalis.

Sensory preconditioning for feeding response in the pond snail, Lymnaea stagnalis.

We demonstrated a sensory preconditioning in the pond snail, Lymnaea stagnalis. An appetitive sucrose solution (a conditioned stimulus: CS1) and weak vibration (another conditioned stimulus: CS2) were first associated, and then the CS2 and an aversive KCl solution (an unconditioned stimulus: UCS) were done. To build the conditioning, two different training procedures, spaced and massed, were examined. After the both training, the sensory preconditioning was built: significantly fewer feeding response to the CS1 became elicited; slower latency to the first bite to the CS1 was induced. No significant differences on the memory retention between these training procedures were found in the sensory preconditioning.

Monosynaptic excitation of motoneurons innervating forelimb muscles following stimulation of the red nucleus in cats.

We examined the rubrospinal projection to forelimb motoneurons (Mns) in cats, in comparison with the corticospinal projection. Under pentobarbital anesthesia, intracellular recordings were made from forelimb Mns following stimulation of the red nucleus (RN) and the cerebral peduncle (CP). Single pulse stimulation of the RN produced excitatory postsynaptic potentials (EPSPs) in the majority of forelimb Mns (58/80, 72%). Segmental latencies of RN-EPSPs were shorter than 1.2 ms in 14 out of 22 Mns in which RN-EPSPs were detected in the C8-T1 segments, and in 3 of 36 Mns in the C6-C7 segments. The results suggest monosynaptic rubro-motoneuronal connections in a substantial portion of forelimb Mns particularly in the C8-T1 segments. There was no evidence suggesting monosynaptic connections between cerebral cortex and forelimb Mns.

Nitric oxide suppresses fictive feeding response in Lymnaea stagnalis.

Fictive feeding activity was monitored in the buccal ganglia of semi-intact preparations of the pond snail, Lymnaea stagnalis, to examine the effects of nitric oxide (NO) released from motoneurons innervating the esophagus on the feeding response. The present results suggest that first; even the low concentration of constitutive NO precisely regulates the feeding rhythm by suppressing high frequency feeding responses; second, that the high concentration of NO released after activation of the feeding central pattern generator following appetitive stimulation of the lips suppresses the feeding rate, resulting in recurrent inhibition. This is the first direct evidence that NO can function to suppress rhythmic activity in the brain.

Enhancement of an inhibitory input to the feeding central pattern generator in Lymnaea stagnalis during conditioned taste-aversion learning.

To study the neuronal mechanism of a conditioned taste-aversion (CTA) learning in the pond snail Lymnaea stagnalis, we examined the synaptic connection between the neuron 1 medial (N1M) cell and the cerebral giant cell (CGC), the former is an interneuron in central pattern generator for the feeding response and the latter is a regulatory neuron to the central pattern generator. Inhibitory postsynaptic potential (IPSP) which was evoked in the N1M cell by activation of the CGC was larger and lasted longer in the conditioned animal than that in the control animal. The electrical properties of the cell body of CGC and the responses of the CGC to the chemosensory inputs were not changed during the CTA learning. These results, together with the previous report indicating the existence of excitatory projection from the N1M cell to the feeding motoneuron, suggest that enhanced IPSP in the N1M cell may underlie the suppression of feeding responses in the Lymnaea CTA learning.

Synaptic inputs of feline rubrospinal neurons from the parietal association cortex, pretectum and medial lemniscus, and their lesion-induced sprouting.

Synaptic inputs of rubrospinal (RN) neurons from the cerebral cortex, pretectal area (PRT), and medial lemniscus (ML) were investigated electrophysiologically in the cat. Stimulation of the ipsilateral parietal association cortex (PASC) and secondary sensory area (SII) produced slow-rising about 3 msec rise time monosynaptic EPSPs which were, in some cases, followed by hyperpolarizations, similar to the sensorimotor cortex (SM)-induced PSPs previously observed. Stimulation of the contralateral cerebral cortex never produced detectable PSPs. Topographical arrangement of PASC-rubral projection was found. Stimulation of the lateral part of PASC induced EPSPs predominantly in RN cells innervating the cervicothoracic spinal segments, while stimulation of the medial part of PASC produced EPSPs predominantly in RN cells innervating the lumbosacral cord. Furthermore, PASC-induced EPSPs were more frequently recorded at the rostral half of RN than at the caudal half. Monosynaptic EPSPs and multisynaptic IPSPs were induced by stimulation of the ipsilateral PRT and ML. PRT- and ML-induced EPSPs had times-to-peak of 1.0 +/- 0.4 msec (mean +/- S.D.) and 1.6 +/- 0.5 msec, respectively, which were intermediate to those of the cerebral peduncle (CP)- and nucleus interpositus of the cerebellum (IP)-induced EPSPs. Furthermore, sensitivity of amplitudes of PRT-induced EPSPs to membrane hyperpolarization was intermediate to those of CP- and IP-EPSPs, and that of ML-induced EPSPs was lower than that of IP-EPSPs. Therefore, it is likely that synapses of PRT and ML fibers are formed between the distal dendrites where CP-rubral synapses terminate and soma where IP-rubral synapses terminate. PASC-induced EPSPs after chronic IP and SM lesions had a new fast-rising component and the effectiveness of ML stimulation to induce the unit spike of RN cells was clearly increased in IP and SM lesioned cats. It was suggested that PASC-rubral fibers sprouted and formed new synapses at the proximal portions of soma-dendritic membranes of RN cells after IP and SM destructions. Collateral fibers to RN cells of the pyramidal tract were also shown to sprout new synapses following IP and ML lesions.

[Laboratory tests on cancer--cytology].

A comparative study was performed on needle aspiration cytology and the histological grading of 34 cases of small liver cancers. According to the Edmondson-Steiner's classification, 9 of the 34 cases were Grade I, 10 cases were a mixture of Grades I and II and 15 were Grade II or a mixture of Grades II and III or Grades I, II and III. The group consisting of Grade II and mixtures of Grades II and III or Grades I, Ii and III were diagnosed by needle aspiration cytology using cytological criteria described previously, but cases consisting of Grade I and mixtures of Grades I and II were very difficult to diagnose because of the relative lack of cellular atypia, and the diagnostic accuracy in this group was low. Retrospectively, ceLlularity, cellular architecture and the nuclear-cytoplasmic ratio were the most valuable cytological findings in the cases of Edmondson-Steiner's Grade I and the mixture of Grades I and II. In addition, some cases of small liver cancers had characteristically large lipid vacuoles which resembled a signet ring cell. Imprint cytology of neuroblastomas is a rapid diagnostic test which is very useful for differential diagnosis of neuroblastoma from other childhood solid tumors. Recently, flow cytometric DNA analysis and the N-myc oncogene have been proposed as important factors in the estimation of the prognosis of neuroblastomas. Three of fore patients with aneuploid DNA under the age of 1.5 years at diagnosis survive, while five of eight patients with diploid DNA over the age of 1.5 years at diagnosis died during a 12 to 120 month follow-up.(ABSTRACT TRUNCATED AT 250 WORDS)

Monosynaptic rubrospinal projections to distal forelimb motoneurons in the cat.

In the present electrophysiological experiments, we examined whether or not there are short-latency excitatory postsynaptic potentials (EPSPs) suggesting monosynaptic rubro- and corticomotoneuronal connections in the cervical spinal segments in cats. Under pentobarbital anesthesia and immobilization with gallamine triethiodide, intracellular recordings were made from forelimb motoneurons (Mns) following stimulation of the red nucleus (RN) and the cerebral peduncle (CP). Mns comprising the musculocutaneous nerve (Musc. Mns), the radial nerve (Rad. Mns), the ulnar nerve (Uln. Mns), and the median nerve (Med. Mns) were identified by their antidromic activation. Single-pulse stimulation of RN produced EPSPs with a mean amplitude of 0.35 mV in the majority of forelimb Mns (88 of 122, 72%). Half of the forelimb Mns (23 of 46) of the C8-T1 segments in which RN-EPSPs were detected by single shocks exhibited segmental latencies of RN-EPSPs shorter than 1.0 ms, while only 3 of 43 (7%) forelimb Mns in the C6-7 segments showed such latencies. These results suggest that rubromotoneuronal connections are, at least in part, monosynaptic for forelimb Mns that innervate in particular the hand and digit muscles. When paired-pulse stimuli (interpulse interval of 3 ms) were applied to RN, the second EPSPs were potentiated moderately (mean amount of temporal facilitation 24%) for EPSPs with short segmental latencies (< 1.0 ms). By contrast, single shocks to CP rarely produced EPSPs in forelimb Mns (9 of 106, 8%). There was no obvious evidence to suggest monosynaptic connections between corticospinal fibers and the forelimb Mns. The present results suggest the presence of preferential direct rubrospinal projection to Mns innervating distal forelimb muscles in the cat.