Preservation of denervated muscle form and function by clenbuterol in a rat model of peripheral nerve injury.

The effects of clenbuterol in preserving the form and function of muscle after unilateral sciatic nerve division and epineural repair were investigated in a rat model. The drug (a beta2-adrenoceptor agonist) was administered daily for six weeks by gastric gavage (10 microg/kg body weight), interrupted every 5 days by a 2 day omission of dosing to avoid drug desensitization. Clenbuterol reduced the loss of wet weight, total protein, muscle fibre cross sectional area and (in part) contractile forces in denervated hindlimb muscles, with most effects lasting until reinnervation. The effects were dependent on muscle type, with slow-twitch oxidative muscle (soleus) and mixed-fibre (gastrocnemius) showing greater sensitivity to the drug than fast-twitch muscle (extensor digitorum longus). Anabolic effects on the contralateral innervated muscles tended to be small. The results suggest a potential for the adjuvant use of selective beta -adrenoceptor agonists in the management of peripheral nerve injuries in humans.

The medial prefrontal cortex plays an important role in the excitation of A10 dopaminergic neurons following intravenous muscimol administration.

Intravenous muscimol administration increases the activity of dopaminergic neurons of the A10 cell group, located in the ventral tegmental area. Evidence suggests that this increase in activity is produced by disinhibition following the inhibition of GABAergic ("non-dopaminergic") cells in the ventral tegmental area. We hypothesized that the activation of A10 cells by muscimol is likely to be at least partly caused by the action of excitatory afferents. To verify this, A10 cells were isolated from ipsilateral afferent sources which utilise excitatory amino acids (which play an important role in the activity of these neurons), using hemisections at the level of the subthalamic nucleus (or just anterior to the subthalamic nucleus), electrolytic lesions of the pedunculopontine tegmental nucleus, or a combination of both. Following hemisections, and hemisections combined with lesions of the pedunculopontine tegmental nucleus, muscimol inhibited rather than excited A10 dopaminergic neurons. The pedunculopontine tegmental nucleus itself appeared to make little intrinsic contribution to muscimol-induced excitation, although the results suggested that part of the excitation which originates in the forebrain may be conducted to A10 cells via the pedunculopontine tegmental nucleus. The source of the effective forebrain excitation was investigated using electrolytic lesions of documented sources of excitatory amino acidergic afferents to the ventral tegmental area: the medial prefrontal cortex, certain nuclei of the amygdalar complex and the lateral habenular nucleus. In the medial prefrontal cortex-lesioned group, muscimol again produced inhibition, an effect qualitatively and quantitatively similar to that in the hemisected groups. Habenular lesions blocked muscimol-induced excitation without producing inhibition, whilst amygdalar lesions produced no significant change in the effects of muscimol. The results suggest that under normal circumstances, an active excitation counteracts and exceeds the direct inhibitory effects of muscimol on the activity of A10 dopaminergic neurons. Furthermore, this activation appears to be produced by the action of excitatory (probably excitatory amino acidergic) afferents arising from the medial prefrontal cortex, and possibly the lateral habenular nucleus. Insofar as the excitation of A10 dopaminergic neurons, which is produced by certain drugs of abuse, and which may play a crucial role in their sustained use, has its basis in excitation following disinhibition, this excitation may provide a novel target for therapeutic intervention in addiction.

D-Amphetamine potentiates muscimol-induced disinhibition of A10 dopaminergic neurons in the rat.

UNLABELLED: Evidence suggests that sensitisation to the behavioural effects of d-amphetamine involves a late-onset (>3 hrs), long-term potentiation (LTP)-like change at medial prefrontal cortex (mPFC)-regulated synapses on A10 dopaminergic (DA) neurons. Since muscimol-induced excitation of A10 DA neurons is dependent on mPFC-regulated afferents, this assay was used to assess whether d-amphetamine enhances the driving of A10 DA neurons by the mPFC, as would be predicted if it resulted in the conditions necessary for LTP. Animals were administered d-amphetamine or saline, 3-4.5 hrs prior to recording. In the acute condition, animals were drug-naïve prior to d-amphetamine, whilst in the challenge condition, animals had previously received d-amphetamine (or saline) each day for 6 days. Recording took place on withdrawal day 2. Muscimol produced significantly less inhibition of A10 DA neurons from animals administered d-amphetamine (rather than saline), but only when d-amphetamine had been chronically administered beforehand (i.e. in the challenge condition). Hence, although the studies fail to provide evidence that acute d-amphetamine administration produces the conditions necessary for LTP, chronic d-amphetamine administration appears to potentiate the impact on A10 DA neurons of mPFC-regulated excitatory activity, thus strengthening the link between this potentiation and the sensitisation process. KEYWORDS: Ventral tegmental area, excitatory amino acids, medial prefrontal cortex, non-DA neurons, synaptic plasticity, behavioural sensitisation.

Stimulation of the pedunculopontine tegmental nucleus in the rat produces burst firing in A9 dopaminergic neurons.

Stimulation of the medial prefrontal cortex in the rat produces events in midbrain dopaminergic neurons which resemble natural bursts, and which are closely time-locked to the stimulation, albeit with a very long latency. As a consequence, we have previously argued that such bursts are polysynaptically generated via more proximal excitatory amino acidergic afferents, arising, for example, from the pedunculopontine tegmental nucleus. In the present study, single-pulse electrical stimulation applied to this nucleus (and other sites in the rostral pons) was found to elicit responses in the majority of substantia nigra (A9) dopaminergic neurons. Responses usually consisted of long-latency, long-duration excitations or inhibition-excitations. Thirty-seven percent of responses (currents combined) elicited by stimulation of the pedunculopontine tegmental nucleus contained time-locked bursts, the bursts being embedded in the long-duration excitatory phases of excitation and inhibition-excitation responses. Stimulation sites located within 0.5 mm of the pedunculopontine tegmental nucleus were also effective at eliciting time-locked bursts (although less so than sites located in the nucleus itself), whereas more distal sites were virtually ineffective. For responses containing time-locked bursts, a higher percentage of stimulations produced a burst when the response was elicited from within the pedunculopontine tegmental nucleus than when it was elicited from outside: the bursts themselves having a very long latency (median of 96.2 ms; shorter than that of medial prefrontal cortex-induced bursts). Finally, although there was no difference in the distribution within the substantia nigra pars compacta of cells which exhibited time-locked bursting and those which did not, stimulation-induced bursts were elicited more frequently in dopaminergic neurons which were classified as "bursting" on the basis of their basal activity. The pedunculopontine tegmental nucleus appears to be a critical locus in the rostral pons for the elicitation of time-locked bursts in A9 dopaminergic neurons. Since time-locked bursts were more often elicited from cells which exhibited bursting under basal conditions, this suggests that rostral pontine sites, in particular the pedunculopontine tegmental nucleus, may play a role in the natural burst activity of dopaminergic neurons. Given that bursts in dopaminergic neurons are generated in response to primary and secondary reinforcers, the projection from the pedunculopontine tegmental nucleus could be one means by which motivationally relevant information (arising, for example, from the medial prefrontal cortex) reaches these cells.

Long-term potentiation at excitatory amino acid synapses on midbrain dopamine neurons.

Evidence suggests that a process analogous to long-term potentiation (LTP) may underlie the enhanced behavioural responses attending chronic administration of amphetamine and cocaine in animals (behavioural sensitization). Augmented excitatory amino acid (EAA)-mediated transmission at the level of midbrain dopamine neurons has been implicated as a change critical to the development of sensitization. Here we provide an initial demonstration that EAA synapses on dopamine neurons can undergo plasticity. Tetanic stimulation of the subthalamic nucleus induced a long-lasting increase (39.2 +/- 10.4%) in the amplitude of excitatory postsynaptic potentials recorded in dopamine neurons of the substantia nigra. This LTP, which did not occur in the presence of NMDA antagonists, may constitute the mechanism that lies at the heart of sensitization.

Differential modulation of voltage-activated conductances by intracellular and extracellular cyclic nucleotides in leech salivary glands.

1. Two-electrode voltage clamp was used to study the effects of adenosine 3':5'-cyclic monophosphate (cyclic AMP) and guanosine 3':5'-cyclic monophosphate (cyclic GMP) on voltage-dependent ion channels in salivary gland cells of the leech, Haementeria ghilianii. 2. Intracellular cyclic AMP specifically blocked delayed rectifier K+ channels. This was shown by use of 3-isobutyl-1-methylxanthine (IBMX, a phosphodiesterase inhibitor), forskolin (an activator of adenylyl cyclase) and intracellular injection of cyclic AMP and its dibutyryl and 8-bromo analogues. Cyclic AMP appeared to be the second messenger for the putative neuroglandular transmitter, 5-hydroxytryptamine. 3. Intracellular injection of cyclic GMP specifically potentiated high-voltage-activated (HVA) Ca2+ current and the effect was mimicked by zaprinast, an inhibitor of cyclic GMP-dependent phosphodiesterase. 4. Extracellularly, cyclic GMP and cyclic AMP specifically decreased the amplitude and increased the rate of inactivation of HVA Ca2+ current. These effects of the cyclic nucleotides are identical to those known for extracellular ATP, which activates a presumed purinoceptor. The pyrimidine nucleotide, UTP, was almost equipotent to ATP (threshold dose < 10(-6) M), indicative of a vertebrate-type nucleotide receptor. However, suramin (5 x 10(-5) M), a non-specific P2-receptor antagonist, failed to block the effects of 5 x 10(-6) M ATP (higher suramin doses could not be reliably tested because of the depolarization and increase in membrane conductance produced by the drug). 5. Activation of the putative purinoceptor by ATP did not affect inward rectifier Na+/K+ current which is known to be potentiated by intracellular cyclic AMP and reduced by intracellular cyclic GMP. 6. The preparation may provide a useful model for study of nucleotide actions, and interactions, in channel modulation. It has technical advantages such as large cells (1200 microns in diameter) which lack intercellular coupling and may be individually dissected for biochemical studies.

Intracellular pH of giant salivary gland cells of the leech Haementeria ghilianii: regulation and effects on secretion.

1. Intracellular pH (pHi) and membrane potential (Em) of giant salivary gland cells of the leech, Haementeria ghilianii, were measured with double-barrelled, neutral-carrier, pH-sensitive microelectrodes. 2. Em was -51 +/- 11.2 mV and pHi was 6.98 +/- 0.1 (mean +/- S.D., N = 41) in Hepes-buffered saline (nominally HCO3(-)-free; extracellular pH, pHe = 7.4). pHi was independent of Em. 3. Amiloride (2 mmol l-1) had no effect on resting pHi or on pHi recovery from an acid load (induced by the NH4+ pre-pulse technique). Removal of external Na+ produced a progressive acidification which was blocked by amiloride, and the drug also slowed the recovery of pHi on reintroduction of Na+. The results indicate the presence of an electroneutral Na+/H+ exchanger whose access to amiloride is competitively blocked by Na+. 4. In certain smaller cells of the gland, which probably form a separate population, removal of external Na+ did not affect pHi, and recovery from an acid load was blocked by amiloride. There may, therefore, be two types of Na+/H+ exchanger, differing in reversibility and sensitivity to amiloride. 5. Recovery of pHi from NH4(+)-induced acid loading was not affected by bicarbonate-buffered saline (2% CO2; 11 mmol l-1 HCO3-) or by addition of the anion-exchange blocker SITS (10(-4) mol l-1). This suggests that there is no significant contribution of a HCO3(-)-dependent transport mechanism to pHi regulation in the gland cells. 6. Removal of external Cl- slowly reduced pHi and there was a transient increase (overshoot) in pHi when Cl- was reintroduced. These effects of Cl- are probably explained by changes in the Na+ gradient. Intracellular Na+ and Cl- activities were measured with ion-selective microelectrodes. 7. Acidification with NH4+ was difficult, probably because of the cells' poor permeability to this ion. Attempts to introduce NH4+ via the Na+ pump or Na+/Cl- transporter were not successful. The H+/K+ ionophore nigericin (1 microgram ml-1), however, produced a rapid and reversible acidification. 8. N-methylmaleimide (0.5-1 mmol l-1), which blocks proton-pumping ATPase, produced a prolonged acidification of almost 1 pH unit, well beyond the level expected for simple equilibration with pHe. The results are consistent with the presence of a vesicular proton pump, acidifying the secretory vesicles which pack the cell body. 9. NH4+ (50 mmol l-1) or trimethylamine (50 mmol l-1) increased pHi and stimulated salivary secretion, while propionate (50 mmol l-1) decreased pHi and stopped secretion.(ABSTRACT TRUNCATED AT 400 WORDS)

Diversity of animal models of aggression: their impact on the putative alcohol/aggression link.

Many attempts have been made to "model" the putative link between alcohol and "aggression" using a variety of situations employing a wide range of animal species. In general, these attempts have proved somewhat disappointing. One reason for this appears to be the fact that the different "aggression" tests tap varied mixtures of offensive, defensive and predatory motivations (i.e., they are not homogenous). Indeed, one can make the claim that the majority of examples of "aggression" associated with alcohol-ingesting humans are inappropriate (nonadaptive) responses whereas many animal models tap adaptive territorial or socially competitive actions. A further problem is that alcohol has very wide-ranging effects on neurophysiology and endocrine functioning. Recent evidence from studies on infra-human animals and clinical situations suggests that some of alcohol's effects on social conflict are expressed by actions in signaling and perception rather than motivation. This aspect is briefly examined in some studies with laboratory mice.

Ethanol-induced enhancement of defensive behavior in different models of murine aggression.

The effects of alcohol on agonistic behavior in mice were studied by introducing an intruder mouse to a resident, alcohol-treated test animal (or saline-injected control). Alcohol (0.1-2.0 g/kg, IP) was administered 20 minutes before testing, and an ethological analysis was made of all behavioral elements shown by the treated animal during a 500-second period. Alcohol did not increase aggression, whether baselines were high, low or experimentally suppressed. Defensive activities, however, were dose-dependently increased, with a threshold dose of 0.5 g/kg or lower in some situations. This suggests that alcohol did not reduce "anxiety" or "fear." Aggression tended to decrease, even with doses as low as 0.5 g/kg, which produced BACs of only 25-40 mg/dl at the start of the testing period. With the highest dose, however, aspects of timidity were still increased after 3 hours, but aggression returned to control level after 1 hour, when the BAC was about 250 mg/dl. In other studies, increased aggression has generally been found only with low alcohol doses. This acute tolerance to the anti-aggressive effect of alcohol reported here suggests the possibility of finding pro-aggressive effects at much higher BACs, perhaps more closely simulating the human situation.

Adriamycin, bleomycin and vincristine chemotherapy with recombinant granulocyte-macrophage colony-stimulating factor in the treatment of AIDS-related Kaposi's sarcoma.

OBJECTIVE: To determine the maximum tolerated dose of granulocyte-macrophage colony-stimulating factor (GM-CSF) that would reduce the severity and duration of neutropenia from combination cytotoxic chemotherapy in the treatment of AIDS-related Kaposi's sarcoma (KS). DESIGN: Phase I, dose escalation. SETTING: Outpatient clinic of a university hospital. PATIENTS: HIV-seropositive patients with advanced KS. INTERVENTIONS: Combination chemotherapy consisting of adriamycin, bleomycin, and vincristine (ABV), with escalating doses of recombinant human GM-CSF (rhGM-CSF). Patients were treated for a median of six cycles (range, between two and seven cycles) of biweekly chemotherapy with GM-CSF administered in divided daily subcutaneous doses on days 2-12. Serum cytokine levels of interleukin (IL)-1 beta, IL-6, and tumor necrosis factor (TNF)-alpha were measured before, during, and after therapy to correlate with response to therapy. RESULTS: A GM-CSF dose of 250 micrograms/m2 was well tolerated, whereas the next dose escalation, of 500 micrograms/m2, was associated with dose-limiting toxicities, including grade 3 fever, fatigue, and diarrhea. GM-CSF produced predictable cyclic increases in granulocytes, allowing for delivery of full-dose chemotherapy on schedule. All patients were HIV-p24-antigen-negative at study entry; no activation of p24 antigenemia was observed after repeat testing. Consistent changes in cytokine levels were not observed. Responses included one complete and three partial responses, and two patients with stable disease parameters. CONCLUSIONS: We conclude that GM-CSF can be administered safely to patients with AIDS-related KS receiving myelosuppressive chemotherapy, resulting in granulocytic response, without up-regulation of HIV p24 antigen levels in serum.

Modulation of inwardly rectifying Na(+)-K+ channels by serotonin and cyclic nucleotides in salivary gland cells of the leech, Haementeria.

The electrically excitable salivary cells of the giant Amazon leech, Haementeria, display a time-dependent inward rectification. Under voltage clamp, hyperpolarizing steps to membrane potentials negative to about -70 mV were associated with the activation of a slow inward current (Ih) which showed no inactivation with time. The time course of activation of Ih was described by a single-exponential function and was strongly voltage dependent. The activation curve of Ih ranged from -72 to -118 mV, with half-activation occurring at -100 mV. Ion-substitution experiments indicated that Ih is carried by both Na+ and K+ ions. 5-Hydroxytryptamine (5-HT) increased the amplitude of Ih and its rate of activation. It also produced a positive shift of the activation curve of the conductance underlying Ih (Gh) without altering the slope factor, thus indicating that the voltage dependence of Ih was modulated by 5-HT. Cs+ blocked both Ih and the 5-HT-potentiated current in a voltage-independent manner, whereas Ba2+ had little effect. It is concluded that 5-HT increases Ih by modulating the inwardly rectifying Na(+)-K+ channels in the salivary cells. The effect of 5-HT may be mediated by an increase in adenylate cyclase activity since Ih was increased by 8-bromo-cyclic AMP and by the phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine. In contrast, Ih was reduced by 8-bromo-cyclic GMP and by zaprinast (an inhibitor of cyclic GMP-sensitive phosphodiesterase). Cyclic GMP itself also reduced Ih, and the effect was specific to the 3',5' form; 2',3'-cyclic GMP was inactive. The results suggest that the inward-rectifier channel may be modulated in opposite directions by cyclic AMP and cyclic GMP.

Rapid co-transport of sodium and chloride ions in giant salivary gland cells of the leech Haementeria ghilianii.

1. Double-barrelled Cl(-)-selective microelectrodes were used to measure the apparent intracellular Cl- activity (aiCl) and membrane potential (Em) of leech salivary gland cells. In standard physiological solution buffered with HEPES (10 mM), intracellular Cl- activity (corrected for interference) was 38 +/- 8 mM (n = 11) compared to a value of 12.8 mM expected for passive Cl- distribution. The mean Em was -49.4 +/- 8.2 mV (n = 21) which was about 27 mV negative to the Cl- equilibrium potential. 2. Removal of external Cl- led to a slow fall in aiCl until a steady-state level of 4-11 mM was reached in 30-60 min. Recovery of aiCl on readdition of external Cl- took only 2-3 min. The uptake followed an exponential time course having a single rate constant of 1.73 +/- 0.1 min-1 (n = 5) whereas the loss appeared to occur in two phases. Changes in external Cl- produced immediate changes in Em which were the opposite of those expected for a high Cl- permeability, i.e. Cl- removal produced an immediate hyperpolarization (3-18 mV) and readdition of Cl- produced a transient depolarization (5-22 mV). 3. The intracellular accumulation of Cl- was dependent on the external Cl- activity. Even when the external Cl- concentration was reduced to 3%, the cells accumulated Cl- against an electrochemical gradient. 4. Cl- accumulation was temperature sensitive (Q10 approximately 2). 5. On removal of external Na+, aiCl fell to a level which was close to that expected for passive distribution. The active reaccumulation of Cl-, after intracellular Cl- depletion, was abolished in the absence of external Na+; aiCl slowly increased to its passive level. Steady-state aiCl or its recovery by Cl(-)-depleted cells was not affected by the absence of K+ in the bathing solution. 6. The reaccumulation of Cl- was not affected by furosemide (1-5 mM), bumetanide (10(-4) M), amiloride (10(-3) M) or 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid (SITS, 10(-4) M). 7. Removal of external Cl- caused a fall in intracellular Na+ activity (aiNa, measured with Na(+)-selective microelectrodes) from 15.9 +/- 6.8 mM (n = 9) to 2.5 +/- 1.3 mM (n = 3). When external Cl- was readded, aiNa rose to 46.5 +/- 6.6 mM (n = 3) before slowly recovering towards its original value. The maximal change in aiNa was 41.7 +/- 4.5 mM (n = 3) and the rate constant for Na+ uptake was 1.8 +/- 0.4 min-1 (n = 3).(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of ethanol on aggression in three inbred strains of mice.

The effects of acutely administered ethanol (0, 0.5 and 1.0 g/kg, IP) were studied in 3 inbred strains of mice during 500 sec dyadic encounters with non-drugged, intruder "standard opponents." The test animals were male C57, DBA and BALB strain mice which had been isolated for 14 or 28 days to promote aggression. The times allocated to 6 broad categories of behaviour were measured from videotape recordings. There was no significant effect of ethanol on threat behaviour or overt aggression, except in 14 day isolates of the DBA strain where aggressive behaviour was reduced by the high dose. This dose increased flight/defensive and freezing behaviour in C57 and BALB mice isolated for 14 days. Ethanol did not affect non-social or social investigative behaviour, except in BALB mice where social behaviour was significantly reduced by the high dose. Blood ethanol levels were similar in DBA and C57 strain mice but were significantly lower in BALB mice.

Effects of naloxone upon the behavioural organisation of specific defeat activities in intruder mice: assessment by cluster analysis.

The present experiment examined the effects of naloxone (0.5, 2.5 and 12.5 mg/kg) upon the responses of male Swiss mice to attack by aggressive male conspecifics in the resident-intruder paradigm by measuring the time spent in broad behavioural categories, frequency of individual acts or postures, and performing a cluster analysis of activities according to their frequency and position within the behavioural sequence. The former two measures detected little naloxone-induced change in behaviour. Cluster analysis revealed changes in behavioural organisation which suggested modification in the motivation and/or function underlying specific defeat behaviour.

Alcohol and social behaviour in group-housed female mice.

Group-housed female mice were injected IP with 0, 0.5, 1.0 or 2.0 g/kg of ethanol. Twenty min after injection, the animal's responses in a neutral cage to a docile male "standard opponent" were videotaped for 500 sec. The tapes were subsequently analysed in terms of the total times allocated by the females to non-social, social/sexual, aggressive and timid activities. The frequency of occurrence of acts and postures associated with the above categories was also determined. Similar females given the same doses were used to assess the blood alcohol concentrations 20 min after injection. Timid behaviour was greatly increased by high doses of ethanol, especially as a consequence of increases in the element 'cringe'; at the low dose 'cringe' was decreased in frequency. Social/sexual behaviour was also influenced in a biphasic manner, low doses increasing and high doses reducing such activity. The drug modified some elements of non-social behaviour in a manner suggesting motor impairment at high doses. There was no evidence that any dose of ethanol induced aggressiveness in these animals. There did not seem to be a simple dose-response effect.

Effects of ethanol on social behaviour directed towards male intruders by lactating Swiss mice.

Ethanol (0.5, 1.0 or 2.0 g/kg) was given intraperitoneally to lactating Swiss strain mice and the animal's response to a docile male intruder was assessed using a videotape analysis that estimated the time that the females allocated to non-social, social/sexual, aggressive and timid categories of response. No evidence was produced in this study that alcohol augmented aggression although conditions were certainly conducive for such an effect. Alcohol does, however, modify aspects of social behaviour in lactating female mice.

Substrate soiled by an unfamiliar conspecific modifies opioid activity in mice placed in novel environments.

Naloxone induces behavioural changes in rodents exposed to novel environments, indicating the involvement of endogenous opioid mechanisms in these situations. The present study investigated whether soiled sawdust substrate from the cage of an unfamiliar, isolated, male conspecific modifies the effect of naloxone (0.5 or 12.5 mg/kg) upon behaviour of mice in an open field test situation. There was little difference between the effects of naloxone upon the frequency of acts or postures shown in the soiled and unsoiled environments. Cluster analysis of the activities according to their position and frequency in behavioural sequences, revealed variations in behavioural organisation in these two situations in control animals, and differential responses to naloxone administration. The data are discussed in terms of an involvement in behaviour of opioid mechanisms which can be modified by non-painful, biologically-relevant, aversive stimuli such as unfamiliar, conspecific-soiled substrates.

Novel preparation for studying excitation-secretion coupling in the isolated single cell.

Cellular mechanisms of secretion in exocrine and endocrine glands are technically difficult to study. We present here a model which offers fundamental advantages for studying excitation-secretion coupling at the level of the isolated single cell. The salivary gland of the leech Haementeria ghilianii possesses a unique combination of unusual properties which greatly facilitate research in this area. Its cells are exceptionally large (up to 1 mm in diameter), clearly visible and easy to penetrate with microelectrodes. They do not form a homogeneous population but consist of five distinct histochemical types, secreting a number of identified products such as the fibrinolytic enzyme hementin (for which there is a sensitive assay). The cells generate overshooting calcium-dependent action potentials up to 90 mV in amplitude and with a duration of 200-1,000 ms. One of their most useful and unusual features is a lack of electrical coupling which means that individual cells can be studied in an intact gland without interference from neighbouring cells.