George Hirst

Role of interstitial cells of Cajal in neural control of gastrointestinal smooth muscles

Specialized cells known as interstitial cells of Cajal (ICC) are distributed in specific locations within the tunica muscularis of the gastrointestinal tract and serve as electrical pacemakers, active propagation pathways for slow waves, and mediators of enteric motor neurotransmission. Recent morphological studies have provided evidence that motor neurotransmission in the gut does not occur through loosely defined synaptic structures between nerves and smooth muscle, but rather via synaptic‐like contacts that exist between varicose nerve terminals and intramuscular ICC (ICC‐IM). ICC‐IM are coupled to smooth muscle cells via gap junctions and electrical responses elicited in ICC are conducted to muscle cells. Electrophysiological studies of the stomach of wild‐type and mutant animals that lack ICC‐IM have provided functional evidence for the importance of ICC in cholinergic and nitrergic motor neurotransmission. The synaptic‐like contacts between nerve terminals and ICC‐IM facilitate rapid diffusion of transmitters to specific receptors on ICC. ICC‐IM also play a role in generating unitary potentials in the stomach that contribute to the excitability of the gastric fundus and antrum.

Propagation of pacemaker activity in the guinea-pig antrum

Cyclical periods of depolarization (slow waves) underlie peristaltic contractions involved in mixing and emptying of contents in the gastric antrum. Slow waves originate from a myenteric network of interstitial cells of Cajal (ICC‐MY). In this study we have visualized the sequence and propagation of Ca2+ transients associated with pacemaker potentials in the ICC network and longitudinal (LM) and circular muscle (CM) layers of the isolated guinea‐pig gastric antrum. Gastric antrum was dissected to reveal the ICC‐MY network, loaded with Fluo‐4 AM and activity was monitored at 37°C. Ca2+ waves propagated throughout the ICC‐MY network at an average velocity of 3.24 ± 0.12 mm s−1 at a frequency of 4.87 ± 0.16 cycles min−1 (n= 4). The propagation of the Ca2+ wave often appeared ‘step‐like’, with separate regions of the network being activated after variable delays. The direction of propagation was highly variable (Δ angle of propagation 44.3 ± 10.9 deg per cycle) and was not confined to the axes of the longitudinal or circular muscle. Ca2+ waves appeared to spread out radially from the site of initiation. The initiating Ca2+ wave in ICC‐MY was correlated to secondary Ca2+ waves in intramuscular interstial cells of Cajal, ICC‐IM, and smooth muscle cells, and the local distortion (contraction) in a field of view. TTX (1 μm) had little effect on slow wave or pacemaker potential activity, but 2‐APB (50 μm) blocked all Ca2+ waves, indicating a pivotal role for intracellular Ca2+ stores. Nicardipine (2 μm) eliminated the Ca2+ transient generated by smooth muscle, but did not affect the fast upstroke associated with ICC‐MY. These results indicate that slow waves follow a sequence of activation, beginning with the ICC‐MY and ICC‐IM network, followed later by a sustained Ca2+ transient in the muscle layers that is responsible for contraction.

Properties of unitary potentials generated by intramuscular interstitial cells of Cajal in the murine and guinea-pig gastric fundus

Intracellular recordings were made from isolated bundles of the circular muscle layer of mouse and guinea‐pig gastric fundus. These preparations displayed an ongoing discharge of membrane noise (unitary potentials), similar to that recorded from similar preparations made from the circular layer of the antrum. Bundles of muscle from the fundus of W/WV mice, which lack intramuscular interstitial cells of Cajal (ICCIM) lacked the discharge of membrane noise observed in wild‐type tissues. When the membrane potential was changed by passing depolarizing or hyperpolarizing current pulses, the discharge of membrane noise was little changed. The membrane noise was unaffected by adding chloride channel blockers; however, agents which buffered the internal concentration of calcium ions reduced the discharge of membrane noise. Treatment of tissues with CCCP, which interferes with the uptake of calcium ions by mitochondria, also reduced the membrane noise and caused membrane hyperpolarization. Similar observations were made on bundles of tissue isolated from the circular layer of the guinea pig antrum. Together the observations indicate that membrane noise is generated by a pathway located in ICCIM. The properties of this pathway appear to vary dramatically within a given organ. The lack of voltage sensitivity of the discharge of membrane noise in the fundus provides a possible explanation for the lack of rhythmic electrical activity in this region of the stomach.

An electrical description of the generation of slow waves in the antrum of the guinea-pig

This paper provides an electrical description of the generation of slow waves in the guinea‐pig gastric antrum. A short segment of a circular smooth muscle bundle with an attached network of myenteric interstitial cells of Cajal (ICC‐MY) and longitudinal muscle sheet was modelled as three electrical compartments with resistive connexions between the ICC‐MY compartment and each of the smooth muscle compartments. The circular smooth muscle layer contains a proportion of intramuscular interstitial cells of Cajal (ICC‐IM), responsible for the regenerative component of the slow wave. Hence the equivalent cell representing the circular muscle layer incorporated a mechanism, modelled as a two stage reaction, which produces an intracellular messenger. The first stage of the reaction is proposed to be activated in a voltage‐dependent manner as described by Hodgkin and Huxley. A similar mechanism was incorporated into the equivalent cell describing the ICC‐MY network. Spontaneous discrete transient depolarizations, termed unitary potentials, are detected in records taken from either bundles of circular smooth muscle containing ICC‐IM or from ICC‐MY. In the simulation the mean rate of discharge of unitary potentials was allowed to vary with the concentration of messenger according to a conventional dose–effect relationship. Such a mechanism, which describes regenerative potentials generated by the circular muscle layer, also simulated the plateau component of the pacemaker potential in the ICC‐MY network. A voltage‐sensitive membrane conductance was included in the ICC‐MY compartment; this was used to describe the primary component of the pacemaker potential. The model generates a range of membrane potential changes with properties similar to those generated by the three cell types present in the intact tissue.

An analysis of inhibitory junction potentials in the guinea‐pig proximal colon

Intracellular recordings were made from either sheets or isolated bundles of the circular muscle layer of guinea‐pig proximal colon and the responses evoked by stimulating inhibitory nerve fibres were analysed. Inhibitory junction potentials (IJPs), evoked by single stimuli, had two components which could be separated on their pharmacological and temporal characteristics and their voltage sensitivities. The initial component, which was abolished by apamin and reduced in amplitude by pyridoxalphosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS), had a brief time course: its amplitude was changed when the external concentration of potassium ions ([K+]o) was changed. The second component of the IJP had a slower onset than the first component, was abolished by l‐nitroarginine (NOLA) and oxadiazolo quinoxalin‐1‐one (ODQ), an inhibitor of soluble guanylate cyclase: its amplitude was little affected by changing [K+]o and was increased when the membrane potential of the circular layer was hyperpolarized. The observations suggest that the initial component of the IJP results from the release of ATP which triggers an increase in membrane conductance to K+ and that the second component results from the release of nitric oxide which suppresses a background inward current.

Stimulated smooth muscle neosphincter in male intrinsic sphincter deficiency: Proof of principle studies in a rabbit model

Intrinsic sphincter deficiency (ISD) causes significant disability and impairment of quality of life despite a range of treatment options. We investigated a novel method to improve sphincter function that does not appear to have been previously attempted, that is, transplantation to create a smooth muscle cuff, that subsequently becomes innervated, around the urethra.