Menachem Hanani

Regeneration of myenteric plexus in the mouse colon after experimental denervation with benzalkonium chloride.

Recent reports suggest a far greater plasticity in nerve tissue than previously believed. As the digestive tract is exposed to a variety of insults, this question is relevant to enteric nerves, but little is known about their ability to recover from damage. To address this problem, we ablated the myenteric plexus of the mouse colon with the detergent benzalkonium chloride (BAC) and followed the ensuing morphologic changes for up to 60 days by using light‐ and electron microscopy. We found that, 2 days after BAC application, the treated area was essentially devoid of intact nerve elements. From day 7, new nerve fibers were observed within the denervated region. This growth progressed until, at days 30–60, newly grown nerve fibers were present in most of this region, and the pattern of muscle innervation was similar to the normal one. At least part of these fibers originated at neurons within intact ganglia surrounding the denervated region. The cross‐sectional area of neurons near the denervated region at day 14 was 52% greater than controls. Glial cells were closely associated with the regenerating nerve fibers. From day 14 onward, we observed undifferentiated cells and differentiating neurons in ganglia surrounding the denervated region, and by day 30, new neurons were present in the myenteric region, along with regenerating nerve fibers. We conclude that the myenteric plexus is endowed with a considerable ability of regeneration and plasticity. The results provide evidence for the presence of stem cells and for an adult neurogenesis in this plexus. J. Comp. Neurol. 462:315–327, 2003.

Satellite glial cells in sensory ganglia: Their possible contribution to inflammatory pain

Neurons in dorsal root ganglia (DRG) are surrounded by an envelope of satellite glial cells (SGCs). Little is known about SGC physiology and their interactions with neurons. In this work, we investigated changes in mouse DRG neurons and SGC following the induction of inflammation in the hind paw by the injection of complete Freunds adjuvant (CFA). The electrophysiological properties of neurons were characterized by intracellular electrodes. Changes in coupling mediated by gap junctions between SGCs were monitored using intracellular injection of the fluorescent dye Lucifer yellow. Pain was assessed with von Frey hairs. We found that two weeks after CFA injection there was a 38% decrease in the threshold for firing an action potential in DRG neurons, consistent with neuronal hyperexcitability. Injection of Lucifer yellow into SGCs revealed that, compared with controls, coupling by gap junctions among SGCs surrounding adjacent neurons increased 2.7-, 3.2-, and 2.5-fold one week, two weeks, and one month, respectively, after CFA injection. In SGCs enveloping neurons that project into the inflamed paw this effect was more enhanced (5.4-fold). Interneuronal coupling was augmented by up to 7% after CFA injection. Pain threshold in the injected paw decreased by 13%, 16%, and 11% compared with controls at one week, two weeks, and one month, respectively, after CFA injection. Intraperitoneal injection of the gap junction blocker carbenoxolone prevented the inflammation-induced decrease in pain threshold. The results show that augmented glial coupling is one of the major events occurring in DRG following inflammation. The elevation in pain threshold after carbenoxolone administration provides indirect support for the idea that augmented intercellular coupling might contribute to chronic pain.

Glial cell plasticity in sensory ganglia induced by nerve damage

Numerous studies have been done on the effect of nerve injury on neurons of sensory ganglia but little is known about the contribution of satellite glial cells (SCs) in these ganglia to post-injury events. We investigated cell-to-cell coupling and ultrastructure of SCs in mouse dorsal root ganglia after nerve injury (axotomy). Under control conditions SCs were mutually coupled, but mainly to other SCs around a given neuron. After axotomy SCs became extensively coupled to SCs that enveloped other neurons, apparently by gap junctions. Serial section electron microscopy showed that after axotomy SC sheaths enveloping neighboring neurons formed connections with each other. Such connections were absent in control ganglia. The number of gap junctions between SCs increased 6.5-fold after axotomy. We propose that axotomy induces growth of perineuronal SC sheaths, leading to contacts between SCs enveloping adjacent neurons and to formation of new gap junctions between SCs. These changes may be an important mode of glial plasticity and can contribute to neuropathic pain.

The effects of axotomy on neurons and satellite glial cells in mouse trigeminal ganglion

&NA; Damage to peripheral nerves induces ectopic firing in sensory neurons, which can contribute to neuropathic pain. As most of the information on this topic is on dorsal root ganglia we decided to examine the influence of infra‐orbital nerve section on cells of murine trigeminal ganglia. We characterized the electrophysiological properties of neurons with intracellular electrodes. Changes in the coupling of satellite glial cells (SGCs) were monitored by intracelluar injection of the fluorescent dye Lucifer yellow. Electrophysiology of SGCs was studied with the patch‐clamp technique. Six to eight days after axotomy, the percentage of neurons that fire spontaneously increased from 1.6 to 12.8%, the membrane depolarized from −51.1 to −45.5 mV, the percentage of cells with spontaneous potential oscillations increased from 19 to 37%, the membrane input resistance decreased from 44.4 to 39.5 M&OHgr;, and the threshold for firing an action potential decreased from 0.61 to 0.42 nA. These changes are consistent with increased neuronal excitability. SGCs were mutually coupled around a given neuron in 21% of the cases, and to SGCs around neighboring neurons in only 4.8% of the cases. After axotomy these values increased to 37.1 and 25.8%, respectively. After axotomy the membrane resistance of SGCs decreased from 101 M&OHgr; in controls to 40 M&OHgr;, possibly due to increased coupling among these cells. We conclude that axotomy affects both neurons and SGCs in the trigeminal ganglion. The increased neuronal excitability and ectopic firing may play a major role in neuropathic pain.

Association of the ecto‐ATPase NTPDase2 with glial cells of the peripheral nervous system

Cellular signaling via extracellular nucleotides appears to play a major role in the functioning of the peripheral nervous system. Information regarding the functional characterization of nucleotide P2 receptors or their expression pattern has been accumulating rapidly; however, very little is known regarding the distribution of ecto‐nucleotidases in the periphery. The extracellular level of nucleotides is controlled by ecto‐nucleotidases, whereby the three membrane‐bound members of the ecto‐nucleoside triphosphate diphosphohydrolase (E‐NTPDase) family are of special functional importance. Using enzyme histochemistry and immunostaining, we demonstrate that NTPDase2 is associated with nonmyelinating Schwann cells of the rat sciatic nerve, whereas NTPDase1 is restricted to blood vessel walls. NTPDase2 immunoreactivity was detected from embryonic day E18 onward, suggesting that immature Schwann cells express the enzyme. With the onset of myelination, NTPDase2 immunoreactivity remained associated solely with nonmyelinating Schwann cells. NTPDase2 was absent from perisynaptic Schwann cells but was associated with fibroblasts covering the endplate at some distance. In addition, NTPDase2 immunoreactivity was associated with the satellite glial cells in dorsal root ganglia and sympathetic ganglia, and with the enteric glia surrounding the cell bodies of ganglionic neurons of the myenteric and the submucous plexus. In contrast to NTPDase1, NTPDase2 preferentially hydrolyzes nucleoside triphosphates over nucleoside diphosphates and thus can act either in inactivating or in producing P2 receptor ligands. Our results suggest that NTPDase2 plays an important role in the control of nucleotide‐mediated activation of peripheral neurons or glia and in the dialogue between these two cell types.

Bidirectional calcium signaling between satellite glial cells and neurons in cultured mouse trigeminal ganglia

Astrocytes communicate with neurons, endothelial and other glial cells through transmission of intercellular calcium signals. Satellite glial cells (SGCs) in sensory ganglia share several properties with astrocytes, but whether this type of communication occurs between SGCs and sensory neurons has not been explored. In the present work we used cultured neurons and SGCs from mouse trigeminal ganglia to address this question. Focal electrical or mechanical stimulation of single neurons in trigeminal ganglion cultures increased intracellular calcium concentration in these cells and triggered calcium elevations in adjacent glial cells. Similar to neurons, SGCs responded to mechanical stimulation with increase in cytosolic calcium that spread to the adjacent neuron and neighboring glial cells. Calcium signaling from SGCs to neurons and among SGCs was diminished in the presence of the broad-spectrum P2 receptor antagonist suramin (50 muM) or in the presence of the gap junction blocker carbenoxolone (100 muM), whereas signaling from neurons to SGCs was reduced by suramin, but not by carbenoxolone. Following induction of submandibular inflammation by Complete Freunds Adjuvant injection, the amplitude of signaling among SGCs and from SGCs to neuron was increased, whereas the amplitude from neuron to SGCs was reduced. These results indicate for the first time the presence of bidirectional calcium signaling between neurons and SGCs in sensory ganglia cultures, which is mediated by the activation of purinergic P2 receptors, and to some extent by gap junctions. Furthermore, the results indicate that not only sensory neurons, but also SGCs release ATP. This form of intercellular calcium signaling likely plays key roles in the modulation of neuronal activity within sensory ganglia in normal and pathological states.

The contribution of satellite glial cells to chemotherapy-induced neuropathic pain

Chemotherapy‐induced peripheral neuropathy is a serious side effect in cancer treatment, a major manifestation being neuropathic pain that can be debilitating and can reduce the quality of life of the patient. Oxaliplatin and taxol are common anti‐cancer drugs that induce neuropathic pain by an unknown mechanism. We tested the hypothesis that satellite glial cells in dorsal root ganglia (DRGs) are altered in chemotherapy‐induced peripheral neuropathy models and contribute to neuropathic pain.

Intercellular coupling of interstitial cells of Cajal in the digestive tract

Interstitial cells of Cajal (ICC) are essential for the normal function of the digestive tract, both as pacemakers and as intermediates between nerves and smooth muscle cells. To perform their functions ICC must be electrically coupled both among themselves and to the muscle layers. This review focuses on the role gap junctions play in coupling ICC to ICC, providing a summary of the published literature as well as a critical appraisal of the data. Most of the experimental evidence for gap junction coupling of ICC networks is indirect, and consists of the ultrastructural observation of gap junctions. Dye coupling studies provide consistent support for the role of gap junctions among ICC of certain types. Physiological evidence in support of this role is scarce. The nature of ICC to smooth muscle coupling is even less certain.

Satellite glial cells in sympathetic and parasympathetic ganglia: In search of function

Glial cells are established as essential for many functions of the central nervous system, and this seems to hold also for glial cells in the peripheral nervous system. The main type of glial cells in most types of peripheral ganglia - sensory, sympathetic, and parasympathetic - is satellite glial cells (SGCs). These cells usually form envelopes around single neurons, which create a distinct functional unit consisting of a neuron and its attending SGCs. This review presents the knowledge on the morphology of SGCs in sympathetic and parasympathetic ganglia, and the (limited) available information on their physiology and pharmacology. It appears that SGCs carry receptors for ATP and can thus respond to the release of this neurotransmitter by the neurons. There is evidence that SGCs have an uptake mechanism for GABA, and possibly other neurotransmitters, which enables them to control the neuronal microenvironment. Damage to post- or preganglionic nerve fibers influences both the ganglionic neurons and the SGCs. One major consequence of postganglionic nerve section is the detachment of preganglionic nerve terminals, resulting in decline of synaptic transmission. It appears that, at least in sympathetic ganglia, SGCs participate in the detachment process, and possibly in the subsequent recovery of the synaptic connections. Unlike sensory neurons, neurons in autonomic ganglia receive synaptic inputs, and SGCs are in very close contact with synaptic boutons. This places the SGCs in a position to influence synaptic transmission and information processing in autonomic ganglia, but this topic requires much further work.

Gap junctions in dorsal root ganglia: Possible contribution to visceral pain

Peripheral injuries can lead to sensitization of neurons in dorsal root ganglia (DRGs), which can contribute to chronic pain. The neurons are sensitized by a combination of physiological and biochemical changes, whose full details are still obscure. Another cellular element in DRGs are satellite glial cells (SGCs), which surround the neurons, but little is known about their role in nociception. We investigated the contribution of SGCs to neuronal sensitization in isolated S1 DRGs from a mouse model of colonic inflammation induced by local application of dinitrosulfonate benzoate (DNBS). Retrograde labeling was used to identify DRG neurons projecting to the colon. Cell‐to‐cell coupling was determined by intracellular dye injection, and the electrical properties of the neurons were studied with intracellular electrodes. Pain behavior was assessed with von‐Frey hairs. The dye injections showed that 10–12 days after DNBS application there was a 6.6‐fold increase in gap junction–mediated coupling between SGCs surrounding adjacent neurons, and this occurred preferentially (another 2‐fold increase) near neurons that project to the colon. Neuron–neuron coupling incidence increased from 0.7% to 12.1% by colonic inflammation. Inflammation led to an augmented neuronal excitability, and to a reduced pain threshold. Gap junction blockers abolished the inflammation‐induced changes in SGCs and neurons, and significantly reversed the pain behavior. We propose that inflammation induces augmented cell coupling in DRGs that contributes to neuronal hyperexcitability, which in turn leads to visceral pain. Gap junction blockers may have potential as analgesic drugs.