Our research teams, to see the effect of Pilocarpine and Dopamine has found promising effect on saliva extraction in ticks that are popular ectoparasite of man and animals. Tick salivation is used to test for various immunological, biochemical, and pharmacological activities. Various chemicals are recognized as stimulating agents to induce saliva secretion.
By Prof Dr Abdullah G Arijo
In the present study two chemicals i.e., pilocarpine and dopamine were used to stimulate the salivary glands for increased saliva secretion associated with time duration. For this purpose, three replicated (partially engorged and fully engorged ticks) experiments were conducted using 5µl of 5% (in methanol) pilocarpine and dopamine. Ticks were collected from Tandojam and adjoining areas from infested body parts of cattle. Host animals included both males and females of all age groups. Pilocarpine or dopamine was pipette onto the scutum of the tick then the glass pipette was pushed into the tick’s mouth part with the pulps remaining outside. Mounted salivating ticks slide was placed inside a dark chamber with high humidity. After one-hour saliva was collected with the help of a pasture pipette bulb and quantified. When saliva extraction was compared between partially fed and fully fed ticks (pooled data) it was seen that in partially fed ticks pilocarpine and dopamine induce tick yield more saliva when compared with untreated control. In fully fed tick data revealed that pilocarpine yielding 2.226µl saliva, which is high as compared to 1.993 µl in partially fed ticks. In the case of dopamine minute’s difference was noted as fully fed ticks produce 1.772µl saliva as compared to 1.705 µl saliva extracted by partially engorged ticks. It could be concluded that pilocarpine is more effective than dopamine to stimulate the salivary glands and increase saliva secretion.
Pakistan is a tropical country and provides favourable environmental and climatic conditions for the development of ticks. Most of the genera and species of ticks are found in Pakistan (Rasul and Akhtar, 1975). Despite these favourable environmental and climatic conditions very little study has been done to interrogate the classification, ecology and prevalence of ticks, which are responsible for infesting the livestock population in Pakistan. Durrani and Kamal, (2008) found the highest prevalence of ticks in district Kasur Pakistan. Hyalomma ticks were found with the highest frequency followed by Boophilus, Haemaphysalis and Rhipicephalus.
Ticks are potential vectors and known to transmit many infectious agents in animals and man such as Salmonella typhimurium, Pasteurella multocida, and Brucella abortus (Jongejan and Uilenberg, 2004). Ticks belong to the phylum Arthropods and class Arachnids. Ticks are mainly divided into two groups: soft ticks (Argasidae) and hard ticks (Ixodidae). Hard ticks feed for a long time on their host, ranging from several days to weeks; the duration of feeding depends on the type of host, stage of the life cycle of ticks and species of ticks. The adult hard ticks can accommodate a large volume of ingested blood in their body which may be from 200 to 600 times more than the un-fed bodyweight of ticks (Sonenshine, 1991). Once the tick had inserted its mouthpart in the body of the host then the pathogens which are present in the tick saliva entered the human or animal body. They exploit the binding site of the ticks. This is called saliva-activated transmission (SAT). It is the indirect transmission of the different pathogen through bio-active tick saliva molecules in the host (Nuttall et al., 2004). The complexity of salivary glands is not astonishing because they secrete various molecules, which act as anticoagulants (Fogcie, 1959), attachment cement, a variety of enzymes (Gill et al., 1986) pharmacological agents (Dickinson et al.,1976) and osmoregulation during feeding (Tatchell, 1969). Additionally, the salivary glands are the main source of paralytic toxins (Ross, 1926) and other components which induce immunological responses to the feeding of ticks (Gill et al., 1986).
The most effective pharmacological stimulator of tick salivary secretion is dopamine, which has been considered as the endogenous signal regulating the salivary gland (Kaufman, 1976; Kaufman, 1977; Mcswain et al., 1992; Schmidt et al., 1982). Other in-vitro and in-vivo pharmacological stimulators, such as norepinephrine and ergot alkaloids, are less active than dopamine and likely act through the dopamine receptors based on the activities of these chemicals on the recombinant tick dopamine D1 receptor (Simo et al., 2011). Another pharmacological stimulator, pilocarpine, is acting through the cholinergic receptor which activates the neural circuit for salivary secretion in the synganglion (the brain of the tick) because it exhibits activity only when the intact synganglion is attached to the salivary gland (Kaufman, 1978; Kaufman and Wong, 1983). The present research was conducted to explore the impact of pilocarpine and dopamine on the collection of tick saliva and to compare the recovery of tick saliva associated with time duration by using the pilocarpine and dopamine.
Ticks used in the present study were identified as Hyalomma anatolicum, a hard tick that may infest a variety of livestock animals’ occasionally even humans. Tick saliva plays a vital role, however, there are certain issues associated with saliva quantity required for experimentation. This study was designed to compare the effect of pilocarpine and dopamine on tick saliva extraction amount. For this purpose, three replicated experiments (with partially fed ticks and fully fed ticks. were conducted using 5µl of 5% pilocarpine and dopamine respectively.
The volume of saliva extraction with15 partially engorges ticks during the first trial in control ticks compared with pilocarpine and dopamine treated tick data revealed that on an average untreated control ticks oozed out 0.613 µl of saliva compared with 1.832 µl in pilocarpine treated ticks and 1.647 µl in dopamine treated Ticks indicating that pilocarpine has relatively high yielding effects in saliva extraction. The second replicated trial for 15 partially engorge ticks conducted on the same protocol and objectives and data may have revealed that untreated controlled tick yielded 0.726 µl of saliva in untreated control ticks whereas the saliva amount increased to 1.935 µl and 1.711 µl in pilocarpine and dopamine treated ticks. To develop a degree of confidence in data and with a view to the reached concreted conclusion using 45 partially engorged tick’s third trial was set. The average value calculated for each group the untreated control tick flush out 0.573 µl of saliva compared to 2.212 µl and 1.758 µl by the ticks treated with pilocarpine and dopamine treated ticks indicating that pilocarpine when compared to dopamine, has a high yielding effect on tick saliva extraction.
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The concentration of saliva is also associated with the post-blood-feeding period. A partially engorged tick relatively produces less amount saliva when compared with fully engorged ticks (Tirloni et al., 2014). For testing these observations another three replicated experiments were conducted using fully engorged ticks.
The average volume of saliva extraction with fully engorge ticks during the fourth trial in control ticks compared with pilocarpine and dopamine data released that untreated control ticks yielded as low as 1.103µl of saliva compared with 2.255µl in pilocarpine treated ticks and 1.891µl in dopamine treated Ticks. This figure indicates that pilocarpine has relatively high yielding effects in saliva extraction. Using the same parameters another trial was conducted and pooled data for 15 fully engorge ticks used in each treatment revealed that untreated controlled tick yielded 0.885 µl of saliva whereas the saliva amount increased to 2.281 µl in pilocarpine and 1.552 µl in dopamine treated ticks. To develop a degree of confidence in data and with a view to reached concreted conclusion using 45 fully engorged ticks sixth trial was set when the average value calculated for each group the untreated control tick flush out 0.743 µl of saliva compared to 2.143 µl and 1.874 µl by the ticks treated with pilocarpine and dopamine treated ticks indicating that pilocarpine when compared to dopamine, has a high yielding effect on tick saliva extraction.
The findings of this work will certainly help research on various aspects of tick saliva and make the way forward into tick research.