Electrical characteristics and responses to jejunal distension of neurons in Remak's juxta-jejunal ganglia of the domestic fowl

Remak's nerve is a ganglionated nerve trunk found only in birds that runs parallel to the gut from the duodenal–jejunal junction to the cloaca. We report the first electrophysiological characterization of these neurons and their responses to gut distension. A segment of chicken jejunum with attached Remak's nerve was pinned in an electrophysiological chamber. Neurons in Remak's ganglia were impaled with microelectrodes. The adjacent segment of gut was distended with fluid. One hundred and thirty neurons were characterized into three electrophysiological classes: (i) tonic neurons (74 %) fired action potentials spontaneously (frequency 3.5 Hz) and continuously (up to 40 Hz) throughout a depolarizing current pulse; (ii) AD neurons (22 %) fired a brief burst of action potentials (1–10), which were followed by a prolonged after-depolarization (AD) of duration 2.8 ± 0.3 s; and (iii) phasic neurons (4 %) fired an initial burst of action potentials followed by an after-hyperpolarization (duration, 520.0 ± 32.0 ms). Tetrodotoxin (1 μM) abolished action potentials in tonic and AD neurons as well as the after-depolarization. Spontaneous fast excitatory postsynaptic potentials (FEPSPs) occurred in all classes of neurons; they were not observed, however, in ganglia isolated from the jejunum. Intracellular injection of biocytin revealed that neurons could be characterized into four morphological classes. Tonic neurons, which had long and extensive dendritic trees, were Remak's Type I, II and IV neurons. AD neurons also comprised Remak's type II neurons. Phasic neurons were Remak's Type III neurons. Most neurons had axons that projected orally along Remak's nerve. Distension of the jejunum evoked FEPSPs and action potentials in tonic neurons, and repetitive bursts of action potentials (1–4) followed by an after-depolarization in AD neurons. All responses to distension were blocked by hexamethonium (300 μM) and tetrodotoxin (1 μM). In conclusion, neurons in Remak's juxta-jejunal nerve appear to regulate gut motility. Three distinct electrophysiological classes of neurons were observed, all of which appear to be activated by distension sensitive cholinergic intestinofugal neurons in the jejunum. Avians, in addition to having prevertebral sympathetic ganglia similar to those found in mammals, have an additional autonomic ganglionated nerve trunk called the intestinal nerve of Remak or Remak's nerve (Remak, 1847). It runs parallel to the gut from the duodenal–jejunal border to the cloaca. Small nerve fibre bundles span the gut and Remak's nerve at frequent intervals along its length (Remak, 1847; Bennett & Malmfors, 1970; Watanabe, 1972; Akester, 1979; Hodgkiss, 1984b; Lunam, 1993; Lunam & Smith, 1996a). Ganglia are regularly spaced in the jejunal and rectal portions of the nerve but are not visible in the ileum (Lunam, 1993; Lunam & Smith, 1996a). Remak's ganglia consist of both catecholamine and non-catecholamine-containing neurons (Costa, 1966; Bennett & Malmfors, 1970; Cantino, Barasa & Guglielmone, 1982; Young, 1990; Lunam, 1991, 1993). The majority of neurons in juxta-jejunal ganglia are catecholaminergic (48–72 %); this proportion decreases anally to approximately 31–46 % in juxta-rectal ganglia (Lunam, 1993). Immunohistochemical studies have demonstrated that these neurons also label for opioids (Epstein & Dahl, 1982; Lunam, 1993) and most non-adrenergic neurons display both opioid and calbindin immunoreactivity (Lunam, 1993). Extracellular electrophysiological studies revealed that post-ganglionic neurons in juxta-jejunal and juxta-rectal ganglia project orally along the nerve trunk (Hodgkiss, 1984a, 1985). We recently injected biocytin into neurons in Remak's ganglia to examine both their morphology and projections (Lunam & Smith, 1996a). Neurons were classified as Remak's Types I, II, III and IV according to their dendritic trees, soma size and shape. Most neurons in the juxta-jejunal ganglia, like many in the juxta-rectal ganglia, projected orally along Remak's nerve. The axons of other neurons in juxta-rectal ganglia could be traced directly into nerve strands bridging the ganglia and the rectum. Differences in the types of neuron and their projections suggested that extrinsic neural circuits controlling gut function may be different in the small and large intestine. Both extracellular electrophysiological and pharmacological studies have demonstrated that Remak's nerve participates in the regulation of gut motility, although the nature of this control is unclear (Bartlet & Hassan, 1971; Kanazawa, Ohashi & Takewaki, 1980; Ishizuka, Komori & Ohashi, 1982; Hodgkiss, 1984a,b; Komori & Ohashi, 1987). Hodgkiss (1984b) demonstrated that Remak's neurons receive cholinergic input from enteric neurons, and proposed that Remak's nerve may be involved in intestinal reflexes, in a manner analogous to those in mammalian prevertebral ganglia (see Janig & McLachlan, 1987; Furness & Bornstein, 1994; Szurszewski & Miller, 1994). Attempts have been made using intracellular electrodes containing biocytin to correlate the electrophysiology of autonomic neurons with their morphology and projections. This approach has been most effective in the enteric nervous system (ENS) where neurons with distinct electrophysiological classes have been shown to have fairly distinct morphologies and projections that are related to function (Bornstein, Furness, Smith & Trussel, 1991; Smith, Bornstein & Furness, 1992; Furness & Bornstein, 1994). In the sympathetic nervous system, Boyd, McLachlan, Keast & Inokuchi (1996) have recently shown that the three distinct electrophysiological classes of neurons (tonic, phasic and long after-hyperpolarizing (LAH)), which comprise postganglionic neurons in prevertebral and paravertebral ganglia, can be separated into three morphological classes according to their soma size, shape and dendritic trees. This correlation between morphology and electrophysiology appears to be less strict in the sympathetic nervous system than for neurons in the ENS. Some neurons with particular electrophysiological characteristics had shapes that overlapped different morphological classes. As far as we are aware, there have been no studies to examine the electrical properties of neurons in ganglia along Remak's nerve. Also, although there have been a number of electrophysiological studies examining the role of sympathetic neurons in extrinsic reflexes controlling colonic motility, there have been few functional studies examining how such neurons control the small intestine (see Szurszewski & Miller, 1994). To address these deficiencies, we have therefore attempted to correlate the electrophysiological and morphological characteristics of neurons in ganglia along Remak's juxta-jejunal nerve, and to determine their electrical responses to distension of the isolated jejunum. We have published preliminary reports of these studies in abstract form (Lunam & Smith, 1994, 1996b).