Immunohistochemical demonstration of cholinergic structures in central ganglia of the slug (Limax maximus, Limax valentianus)

Immunohistochemical techniques were used to study the distribution of cholinergic neurons containing choline acetyltransferase of the common type (cChAT), the synthetic enzyme of acetylcholine, in the central nervous system of the slug Limax maximus and Limax valentianus. Because the antiserum applied here was raised against a recombinant protein encoded by exons 7 and 8 of the rat gene for ChAT, three methods were used in order to validate antibody specificity for the Limax counterpart enzyme. Western blot combined with ChAT activity assay following native gel electrophoresis and immunoprecipitation analysis both indicated that immunoreactive Limax brain molecules were capable of synthesizing acetylcholine. Western blot after denatured gel electrophoresis of Limax brain extracts revealed a single band of about 67kDa. All findings obtained with these three methods clearly indicated that the antiserum effectively recognized Limax cChAT. 1400 neuronal cell bodies positive for cChAT, mainly small to medium-sized, were found in various brain regions in the buccal, cerebral, pleural, parietal, visceral and pedal ganglia. cChAT immunoreactive nerve fibers were distributed extensively in the neuropil, connectives and commissures of these central ganglia. The map of cChAT-positive cells provided here are valuable for understanding the cholinergic mechanism in the slug brain, as well as giving an important hint to clarifying the mechanisms of learning and memory in higher vertebrates including humans.     

Role of EGR1 in hippocampal synaptic enhancement induced by tetanic stimulation and amputation

Hippocampal neurons fire spikes when an animal is at a particular location or performs certain behaviors in a particular place, providing a cellular basis for hippocampal involvement in spatial learning and memory. In a natural environment, spatial memory is often associated with potentially dangerous sensory experiences such as noxious or painful stimuli. The central sites for such pain-associated memory or plasticity have not been identified. Here we present evidence that excitatory glutamatergic synapses within the CA1 region of the hippocampus may play a role in storing pain-related information. Peripheral noxious stimulation induced excitatory postsynaptic potentials (EPSPs) in CA1 pyramidal cells in anesthetized animals. Tissue/nerve injury caused a rapid increase in the level of the immediate-early gene product Egr1 (also called NGFI-A, Krox24, or zif/268) in hippocampal CA1 neurons. In parallel, synaptic potentiation induced by a single tetanic stimulation (100 Hz for 1 s) was enhanced after the injury. This enhancement of synaptic potentiation was absent in mice lacking Egr1. Our data suggest that Egr1 may act as an important regulator of pain-related synaptic plasticity within the hippocampus.     

In vivo and in vitro patch-clamp recording analysis of the process of sensory transmission in the spinal cord and sensory cortex

Following the integration and modification of the sensory inputs in the spinal cord, the information is transmitted to the primary sensory cortex where the integrated information is further processed and perceived. Processing of the sensory information in the spinal cord has been intensively investigated. However, the mechanisms of how the inputs are processed in the cortex are still unclear. To know the correlation of the sensory processing in the dorsal horn and cortex, in vivo and in vitro patch-clamp recordings were made from rat dorsal horn and sensory cortex. Although dorsal horn neurons showed spontaneous and evoked EPSCs by noxious and non-noxious stimuli, most somatosensory neurons located at 100 to 1000 microm from the surface of the cortex exhibited an oscillatory activity and received synaptic inputs from non-noxious but not noxious receptors. These observations suggest that the synaptic responses in cortical neurons are processed in a more complex manner; and this may be due to the reciprocal synaptic connection between thalamus and cortex.     

Oscillations and gaseous oxides in invertebrate olfaction

Olfactory systems combine an extraordinary molecular sensitivity with robust synaptic plasticity. Central neuronal circuits that perform pattern recognition in olfaction typically discriminate between hundreds of molecular species and form associations between odor onsets and behavioral contingencies that can last a lifetime. Two design features in the olfactory system of the terrestrial mollusk Limax maximus may be common elements of olfactory systems that display the twin features of broad molecular sensitivity and rapid odor learning: spatially coherent oscillations in the second-order circuitry that receives sensory input; and involvement of the interneuronal messengers nitric oxide (NO) and carbon monoxide (CO) in sensory responses and circuit dynamics of the oscillating olfactory network. The principal odor processing center in Limax, the procerebrum (PC) of the cerebral ganglion, contains on the order of 10⁵ local interneurons and receives both direct and processed input from olfactory receptors. Field potential recordings in the PC show an oscillation at approximately 0.7 Hz that is altered by odor input. Optical recordings of voltage changes in local regions of the PC show waves of depolarization that originate at the distal pole and propagate to the base of the PC. Weak odor stimulation transiently switches PC activity from a propagating mode to a spatially uniform mode. The field potential oscillation in the PC lobe depends on intercellular communication via NO, based on opposing effects of reagents that decrease or increase NO levels in the PC. Inhibition of NO synthase slows the field potential oscillation, while application of exogenous NO increases the oscillation frequency. A role for CO in PC dynamics is suggested by experiments in which CO liberation increases the PC oscillation frequency. These design features of the Limax PC lobe odor processing circuitry may relate to synaptic plasticity that subserves both connection of new receptors throughout the life of the slug and its highly developed odor learning ability. © 1996 John Wiley & Sons, Inc.     

Differential expression of the klf6 tumor suppressor gene upon cell damaging treatments in cancer cells

The mammalian Krüppel-like factor 6 (KLF6) is involved in critical roles such as growth-related signal transduction, cell proliferation and differentiation, development, apoptosis and angiogenesis. Also, KLF6 appears to be an emerging key factor during cancer development and progression. Its expression is thoroughly regulated by several cell-damaging stimuli. DNA damaging agents at lethal concentrations induce a p53-independent down-regulation of the klf6 gene. To investigate the impact of external stimuli on human klf6 gene expression, its mRNA level was analyzed using a cancer cell line profiling array system, consisting in an assortment of immobilized cDNAs from multiple cell lines treated with several cell-damaging agents at growth inhibitory concentrations (IC(50)). Cell-damaging agents affected the klf6 expression in 62% of the cDNA samples, though the expression pattern was not dependent on the cell origin type. Interestingly, significant differences (p<0.0001) in KLF6 mRNA levels were observed depending on the cellular p53 status upon cell damage. KLF6 expression was significantly increased in 63% of p53-deficient cells (122/195). Conversely, KLF6 mRNA level decreased nearly 4 fold in more than 70% of p53+/+ cells. In addition, klf6 gene promoter activity was down-regulated by DNA damaging agents in cells expressing the functional p53 protein whereas it was moderately increased in the absence of functional p53. Consistent results were obtained for the endogenous KLF6 protein level. Results indicate that human klf6 gene expression is responsive to external cell damage mediated by IC(50) concentrations of physical and chemical stimuli in a p53-dependent manner. Most of these agents are frequently used in cancer therapy. Induction of klf6 expression in the absence of functional p53 directly correlates with cell death triggered by these compounds, whereas it is down-regulated in p53+/+ cells. Hence, klf6 expression level could represent a valuable marker for the efficiency of cell death upon cancer treatment.     

All in a sniff: olfaction as a model for active sensing

Sensation is an active process involving the sampling and central processing of external stimuli selectively in space and time. Olfaction in particular depends strongly on active sensing due to the fact that-at least in mammals-inhalation of air into the nasal cavity is required for odor detection. This seemingly simple first step in odor sensation profoundly shapes nearly all aspects of olfactory system function, from the distribution of odorant receptors to the functional organization of central processing to the perception of odors. The dependence of olfaction on inhalation also allows for profound modulation of olfactory processing by changes in odor sampling strategies in coordination with attentional state and sensory demands. This review discusses the role of active sensing in shaping olfactory system function at multiple levels and draws parallels with other sensory modalities to highlight the importance of an active sensing perspective in understanding how sensory systems work in the behaving animal.