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The full chemical name of ostarine is 2s 4-cyanophenoxy -n-[4-cyano trifluoromethyl phenyl]hydroxymethyl-propanamide. Ostarine mk is a sarm developed by gtx for the prevention and treatment of muscle wasting. It may eventually be a medical prescription for the prevention of cachexia, atrophy and sarcopenia as well as for hormone or testosterone replacement therapy. The reflex labs advanced muscle stack is a powerful sarm stack for building muscle. This advanced sarms stack including ligandrol lgd, ostarine mk, ibutamoren mk, and testolone rad rapidly accelerate the otherwise time-consuming bulking process.

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Take an average cycle length of two months, you just take three capsules a day before you eat, cardarine daily dose. Even if you do continue steroid use, you also risk losing fertility, making it difficult to have kids in the future. Besides the risk of infertility, if you continue to take anabolic steroids into your mid 20's you also risk disrupting your body's own natural hormone cycle, causing problems with your pituitary gland and testosterone production, cardarine daily dose.

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Nitric oxide is now the new secret weapon used by athletes and bodybuilders to improve performance, cardarine daily dose. Cardarine daily dose, cardarine daily dose This will help you train longer, speed recovery and feed your muscles which leads to more muscle being built even faster.

For this stack, they recommend you take the whey protein after workouts or mid-morning, take the clean gainer after workouts or before bed, take the BCAA's during your workout and the creatine after your workout. Cost of the stack is more than the supplements individually Too focused on protein, could have used a good pre-workout Uses artificial flavors, cardarine daily dose. Where to buy crazy bulk dbal, where to buy testo max A good ostarine cardarine stack will last between 8 to 12 weeks.

Use a normal 20mg on top of 25mg of ostarine daily. Cardarine and stenabolic stack. Stenbolic sr is a fantastic sarm that stacks well with gw This combo is best used to promote fat loss. To stack with sr, use 20mg with 25mg of sr daily. Your doses should be anywhere between mgs a day i took 20mgs daily. I recommend a nice 12 week cycle to get the full benefits. If you are going to run additional cycles take 6 week breaks in-between cycles.

The half-life for cardarine is about 24 hours, so you only need to dose it once a day. Gw cardarine potential risks. Gsk abandoned further development of the drug in because testing on rats showed prolonged use caused cancer to develop in several organs. My personal conclusion is that this does not call into question the original studies that show the incidence of cancer, but reaffirms that length of time running this compound can not be ignored.

Cardarine is usually taken in dosages between 5 mg and 10 mg, but dosages as high as 20 milligrams daily have been shown to be safe in studies. Stenabolic, or sr , is usually taken in dosages ranging from 10 mg to 20 mg per day, although dosages as high as 30 mg have shown to be safe. The recommended dosage of cardarine: cycles after conducting dosage trials, the correct dosage of cardarine to be taken is found to be 10 mg to 20 mg per day, for about 8 weeks.

Just curious how important the timing of your dose of cardarine is for best results? Does it make much difference or can you get the desired results just taking a daily dose regardless of timing? Guidelines for dosing cardarine the recommended daily dose of gw is between mgs. However, if you really want to blast the fat off go for the 20mg per day quota. The daily dosage of cardarine is between 10mg and 20mg. However, if you want to blast the fat off, 20 mg works better.

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The pre-chemo blood tests we were told keep an eye on vital organs reactions so that situations can be monitored. Wishing you all the best with your treatment. One thing the Steroids for anti sickness do is knock my blood sugar for six I have to double my insulin type two diabetic for a couple of days until I can cut out the Dexamethosone! Skip to main content. Post to forum. Search Search forum. Do you have a cancer chat password?

Yes, I have a password. Remember me. Sign in. I would be happy to receive news and updates from Cancer Chat. Create new account. Leave this field blank. Prolonged use of steroids can lead to serious health problems. Insulin is not a steroid but it still makes its way into the steroids list because of its importance and uses in the anabolic growth of muscles. This peptide hormone is similar to hgh as it helps in the building of muscle tissue from the amino acids entering the cells.

Anabolic steroids are synthetic versions of the male hormone testosterone. They have two main effects on the human body; an anabolic, muscle-building, effect and an androgenic, or masculinizing. The anabolic steroids are misused by bodybuilders and athletes for unnatural types of testosterone, generally, men and women develop testosterone naturally.

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Anabolic steroids affect the onset of puberty, the growth of the clitoris in females and the penis in male children does not affect the size of the penis in adults , increased the size of the vocal chords and deepening of the voice, increased body hair, and premature baldness in people predisposed to it. The anabolic steroids control act of placed anabolic steroids into schedule iii of the controlled substances act csa as of february 27, Mk muscle gain, anabolic steroids 1 cycle Anabolic steroids list, order anabolic steroids online visa card.

The muscle-building chemical in Deca Durabolin has similar properties to those of testosterone, meaning it does have the potential to promote muscle growth and cause side effects linked to the consumption of anabolic steroid, anabolic steroids list.

Unlike Deca Durabolin, which has to be injected for efficacy, DecaDuro is formulated in capsule form. DecaDuro promotes nitrogen retention, protein synthesis, and red blood cell production. Nitrogen is one of the components of amino acids, the building blocks of all protein DecaDuro improves the body's ability to retain more nutrients, so there are more building blocks of proteins to metabolize and transform into muscle gain. Deca durabolin nandrolone decanoate D-Bal improved muscle mass growth Enhance the muscle recovery process Safer than Dianabol Improves nitrogen retention Delivered globally, anabolic steroids list.

Anabolic steroids list, price buy legal anabolic steroid cycle. Thus, ALT and AST liver values will not rise dramatically, ensuring that your liver will remain healthy and protected, anabolic steroids sustanon Ostarine mk ostarine mk is very similar to the anabolic compound dvar. Like dvar, mk is designed to reduce body fat, but it can simultaneously increase lean body mass as well.

Notably, ostarine has the best muscle tissue healing ability than all other sarms too. Ostarine is a mild sarm that is known for its muscle-building and performing-enhancing effects. It has been reported that men can expect to gain pounds of muscle per 8-week cycle when taking 25 mg per day of mk If your goal is to lose weight while taking ostarine for a cutting cycle, it is advised that men take mg per day for a week cycle, while women take 10 mg per day for a week cycle.

Gaining lean muscle because ostarine was created to help combat muscle wasting diseases, it has great muscle building abilities. Most people think enobosarm mk can only be used when cutting bodyfat, that is not true. Ostarine mk review — shocking truths you need to know! When using mk in a dose of 30mg a day, you can expect gains of kg lb within 10 weeks.

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Reduced adrenal function adrenal insufficiency. This potentially life-threatening condition can happen when you stop taking oral corticosteroid medicines and start using inhaled corticosteroid medicines such as QVAR RediHaler. Tell your healthcare provider right away about any signs and symptoms of adrenal insufficiency such as: feeling tired or exhausted fatigue ; lack of energy; low blood pressure hypotension ; dizziness or feeling faint; nausea and vomiting; or weakness.

Immune system effects and a higher chance for infections. Tell your healthcare provider about any signs or symptoms of infection such as: fever, chills, pain, feeling tired, body aches, nausea, or vomiting. Always have a rescue inhaler with you to treat sudden wheezing.

Serious allergic reactions. Stop using QVAR RediHaler and call your healthcare provider or get emergency medical help right away if you get any of the following: hives; swelling of your lips, tongue, or face; rash; or breathing problems. Slowed growth in children. Lower bone density. This may be a problem for people who already have a higher chance for low bone density osteoporosis. Eye problems. If you have had glaucoma, cataracts or blurred vision in the past, you should have regular eye exams while using QVAR RediHaler.

Common side effects of QVAR RediHaler include: yeast infection in the mouth oral candidiasis ; cold symptoms upper respiratory tract infection ; pain in the throat oropharyngeal pain ; pain or swelling in your nose and throat nasopharyngitis ; sinus irritation sinusitis ; and hay fever allergic rhinitis These are not all the possible side effects of QVAR RediHaler.

Call your doctor for medical advice about side effects. Approved Use QVAR RediHaler Inhalation Aerosol is a breath-actuated inhaled prescription medicine used as a maintenance treatment for the prevention and control of asthma in people 4 years of age and older. One other prominent feature those using this product will benefit from is that of resistance to the 3-hydroxysteroid dehydrogenase enzyme.

This enzyme, present in all muscle tissues, is responsible for channeling the DHT hormone for non-anabolic purposes throughout the system. It does this in a threefold manner; first, by reducing glucocorticoid hormones present in the system that have been extensively linked with the breakdown of muscle tissue. All of the above information primarily regards what kind of effect this product will have within the system, but anavar also features another modification that affects its general practicality.

This would usually take place as a result of direct intervention via the hepatic metabolism. Home Anavar FAQs. Get Your Free Anavar Cycle. Anavar results specifically will vary depending on how disciplined you are nutritionally and physically in regards to adhering to stringent training principles, but ultimately you can expect to reap a wealth of positive rewards.

Anavar increases your lean muscle mass and improves bone density. Anavar helps you to burn fat, both subcutaneous and visceral fat.

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Hope you will not experience too many problems though always wise to tell your chemo team as and when anything arises so they can try and help. The pre-chemo blood tests we were told keep an eye on vital organs reactions so that situations can be monitored. Wishing you all the best with your treatment.

One thing the Steroids for anti sickness do is knock my blood sugar for six I have to double my insulin type two diabetic for a couple of days until I can cut out the Dexamethosone! Skip to main content. Post to forum. Search Search forum. Do you have a cancer chat password? Yes, I have a password. Remember me. Sign in.

I would be happy to receive news and updates from Cancer Chat. Create new account. Leave this field blank. Already a member? Sign in now. Further, we found transcripts predicting genes for pheromone odorant receptor, gustatory receptor, novel GPCR messages, ecdysone nuclear receptor, JH esterase binding protein, steroidogenic activating protein, chitin synthase, chitinase, and other genes of interest.

Also found were transcripts predicting genes for spermatogenesis-associated protein, major sperm protein, spermidine oxidase and spermidine synthase, genes not normally expressed in the female CNS of other invertebrates. The diversity of messages predicting important genes identified in this study offers a valuable resource useful for understanding how the tick synganglion regulates important physiological functions.

This is an open-access article distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: Funding for the laboratory of D. The funders had no role in study design, data collection and analysis, decision to publish or preparation of the manuscript. Competing interests: One of the co-authors, B.

There are no patents, products in development or marketed products to declare. Ticks are obligate blood-feeding ectoparasites that serve as vectors of the causative agents of many important diseases affecting humans and animals, e. The black-legged tick, Ixodes scapularis , is one of the most important vectors of infectious diseases to humans and animals throughout large areas of the United States and Canada [1] , [3] , [4].

Lyme disease is the most commonly reported vector-borne disease in the northern temperate zone regions of the northern hemisphere [5] , [6] with 24, confirmed cases in the United States in [7]. Despite the many zoonotic diseases caused by tick-borne pathogens, control of ticks is still largely dependent on the use of chemical acaricides, with traditional acaricide targets being primarily neurologically-based [8].

The central nervous system CNS in ticks is composed of a single mass called the synganglion [9]. Despite the fact that most acaricides target the nervous system, relatively little is known about tick neurobiology and how blood-feeding and mating affect gene expression in the synganglion. Acaricide resistance has become widespread around the world causing a need for new acaricide targets or alternative methods of tick control [10]. Continued dependence on traditional chemical methods is becoming increasingly problematic in the face of ever greater demand for environmental protection and fear of chemical poisoning.

To search for alternatives to chemical control, a better understanding of the neurologic processes that control basic tick physiological processes is needed. Ixodid ticks must gorge on blood, often increasing fold in body size, in order to stimulate tissue development, ecdysis, mating and reproduction [9].

Precisely how the tick's nervous system regulates these biological processes is largely unknown. During feeding, some synganglion cells, especially the neurosecretory cells, increase in size and accumulate neurosecretory substances [11] , [12]. Neurohormones and neurotransmitters play key roles in tick development and physiology. Work has been conducted examining their occurrence in selected tissues in ticks such as the hemocytes [13] , midgut [14] , ovaries [13] , and salivary glands [2] , [13] , [15] — [20].

However, much of the work on neurotransmitters focuses on the dopaminergic system in the salivary glands reviewed by [21] — [23]. Much less is known about the transcribed genes in the tick synganglia, largely because of the difficulty in extracting sufficient amounts of tissue.

Advances in sequencing technology now allow researchers to rapidly obtain large amounts of data from a small amount of tissue. Recent work by Simo et al. However, the molecular basis for understanding just how these complex neurohormone-controlled networks regulate tick physiological functions remains to be determined.

To this end, a global search to identify and characterize the numerous molecules expressed in the synganglion is needed. Transcriptomics offers an excellent tool to approach this problem [25]. Using pyrosequencing, Bissinger et al. Previous studies by Donohue et al. Transcriptome analysis of the synganglion of the brown dog tick, Rhipicephalus sanguineus , was done with cDNA library construction in phage resistant Escherichia coli.

However, this method yielded a total of only ESTs sequenced, with only remaining after removal of vector contamination that could be clustered into unique transcripts [20]. Despite these few studies, there is a paucity of information about the neurobiology of ticks.

Along with improvements in assembly technology these advances have greatly enhanced the value of transcriptomes for the study of messages encoding for diverse genes in selected tissues or whole organisms [31] , even in organs as small as the tick synganglion.

Transcriptomes provide an opportunity to examine at high resolution the entire repertoire of mRNA molecules expressed in a particular organ or tissue at a particular moment in time. Transcriptomes have proven useful for the study of gene expression of many arthropods, including mosquitoes [32] , bedbugs [33] and other arthropods, as well as selected organs, such as the transcriptome of the midgut of female D.

Here we present the first transcriptome of the synganglion of the black legged tick, I. We conducted BLAST matching against the published conspecific genome and gene sequence alignments which we believe are likely to increase the reliability of gene annotations in the relevant transcriptomes. In this report, we compare the success of these two different technologies in identifying the many messages predicting genes of interest for our understanding of synganglion function in this important tick species.

We believe that the assembled, annotated transcriptomes described in this report will provide an invaluable resource for future studies of the role of the genes involved in neurologic functioning of the tick nervous system. Black-legged ticks, I. Adult ticks were confined within plastic capsules attached to New Zealand white rabbits, Oryctolagus cuniculus , and allowed to feed to repletion and oviposit.

Larvae and nymphs were allowed to feed on albino mice, Mus musculus. All use of animals in this study was carried out in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health. Adult I. Three different RNA samples were prepared, 1 5.

San Diego, CA. A minimum of 1. Following PCR amplification, adapters were included for sequencing with paired ends. For pyrosequencing, the total RNA was thawed and mRNA isolated from each of 3 female synganglion samples, using an Oligotex mRNA isolation kit according to the manufacturer's recommendations.

The samples were evaluated using the Bioanalyzer as described above to insure that they met the minimum requirements for sequencing. An additional 5 reactions were carried out with 20 cycles the optimized number of cycles to produce sufficient quantities of cDNA for preparation of the library. Following PCR, the contents from the different samples were combined into a single sample and the cDNA was purified using a PCR purification kit Qiagen according to the manufacturer's recommendations.

The only deviation from the protocol was that DNA-positive beads were enriched after emulsification PCR in order to increase the number of reads collected during titration. Enrichment was done so that only beads containing DNA were loaded and the data generated during titration sequencing could also be used in the assembly of contiguous sequences.

Enrichment of DNA-positive beads was completed exactly as described by Margulies et al. We first trimmed the sequences of ambiguous bases, low quality base calls, and by removing any latent primer or adapter sequences.

Transcripts were identified annotated by BLAST [37] against the GenBank nr and EST databases at an E-value of E or lower for the Illumina assemblies and E for although with few exceptions, only matches E or lower were accepted for inclusion in the data tables. Additional evidence to support the annotation of transcripts of interest in the different transcripts was done using alignments with conspecific genes or other arthropod genes.

The version described in this paper is the first version, GBBN The transcriptome was deposited in the TSA and will be published pending curation. The results of Illumina and sequencing for the three different samples are shown in Table 1. For sample Il-1 mixed females , sequencing yielded a total of 34,, raw reads for a total read length of 2,,, Almost all reads were matched 34,, The average read length matched reads was 68 bp, and included a total length of 2,,, bp.

Following vector trimming, the raw reads were assembled to a total of 41, transcripts with an average length of bp, comprising 19,, bp. For sample Il-2 part-fed females , sequencing yielded a total of ,, raw reads. The average length was 72 bp and included a total of 8,,, bp. However, less than half were matched reads 43,, , comprising only 3,,, bp.

Following vector trimming, the raw reads were assembled to a total of 30, transcripts with an average length of bp, comprising 20,, bp. For sample part-fed females , sequencing yielded a total of , raw reads. The average length was bp and included a total length of ,, bp. Following vector trimming, the raw reads were assembled to a total 20, transcripts, with an average length of bp, comprising 10,, bp.

BLAST matching of the transcript files showed that almost all of the top hits matched sequences from the I. Approximately 17, sequences matched similar sequences from I. Overall, there was little difference in the GO assignments in these transcriptomes, although there were 38 transcripts assigned to reproduction in Il-1 mixed females as compared to only 8 transcripts in sample Il-2 part-fed females versus 85 transcripts in sample Categories considered of greatest interest for regulating synganglion biological activity, development and reproduction in the 3 transcriptomes are shown in Table 2 BioPro Level 2.

For BioPro level 3, there was little difference between samples Il-2 and GO term searching success for these samples was comparable to studies of transcriptomes of other tick tissues sequenced by pyrosequencing, e. The GO term assignments for biological processes at level 3 are shown in Figures 3 , 4 , 5 and selected categories of interest in Table 2 BioProcesses Level 3. Many more categories regulating synganglion biological activity were found than were identified in BioPro level 2, including biological regulation, oxidation-reduction, cellular response to stimulus, cell to cell and other signaling types, cellular developmental processes, hormone metabolic processes, immune response and reproduction.

Of special interest for sensory perception was the number of messages for responses to external, abiotic and chemical stimuli, representing as much as 1. Transcripts matching genes involved in cell to cell signaling, signaling processes, cell to cell communication and signaling pathways represented only 0. This figure shows the 56 GO term assignments categorized in this transcriptome 11, transcripts.

This figure shows the 84 GO term assignments categorized in this transcriptome 23, transcripts. This figure shows 81 GO term assignments categorized in this transcriptome 14, transcripts. Comparison of the I. Whether this reflects true differences in expression or is a function of the larger proportion of genes mapped in I. In addition to biological functions, GO term searching also examined the numerous GO categories by molecular function.

At molecular level 3, GO term searching of the transcriptome for sample Il-1 showed 34 categories with a total of 8, genes, or The most abundant categories were hydrolase activity 1, , transferase activity 1, , ion binding , nucleic acid binding , nucleotide binding , protein binding , oxireductase activity , and signal transducer activity GO term searching of the transcriptome for sample Il-2 showed 44 categories with a total of 17, genes, or The most abundant categories were hydrolase activity 2, , transferase activity 2, , ion binding 1, , nucleic acid binding 1, , protein binding 1, , nucleotide binding 1, , oxireductase activity , and signal transducer activity Of special interest for synganglion regulatory activity was neurotransmitter binding 23 genes and kinase regulatory activity 18 genes highlighted in yellow.

In contrast, although the transcriptome for sample showed 58 categories, the total number of genes mapped was only 2, genes, or The most abundant categories were hydrolase activity , nucleotide binding , transferase activity , protein binding , ion binding , oxireductase activity , nucleic acid binding , nucleoside binding and signal transducer activity Little evidence of neurotransmitter receptors or neuropeptides was found at this level, namely neurotransmitter receptor binding with 3 genes and peptide binding with 10 genes.

However, others may have been present but incorporated into broader categories. This figure shows the 34 GO assignments categorized in this transcriptome 8, transcripts. This figure shows the 44 GO assignments categorized in this transcriptome 17, transcripts. This figure shows the 58 GO assignments categorized for this transcriptome 2, transcripts.

Table 2 shows the contrasts for 19 GO molecular categories of interest for the three different transcriptomes as percentages of the total searched genes. Little difference was noted in the percentages of gene messages in these GO categories among the three different transcriptomes. Tables 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 compare the three different transcriptomes with respect to the numbers of transcribed genes i. Transcript frequency indicates the number of transcripts that predicted the same gene identified in GenBank and should not be misconstrued as a measure of gene expression.

The number of true neuropeptides in ticks is uncertain. Estimates of expressed neuropeptides range from as few as 20, characterized by proteomic methods [29] to as many as 80, characterized by in silico searches of publicly accessible EST databases [28].

In the most recent estimate, genes for 56 neuropeptides have been identified in I. Of the 56 neuropeptide genes believed to occur in this tick Hill et al. Comparing our findings with the transcriptome of D. The actual number of mature peptides is likely somewhat greater since in some cases several neuropeptides can be processed via post-translational modifications from a prepropeptide, e.

We also found transcripts encoding for 7 other neuropeptides, namely, allatotropin, corticotropin-releasing factor CRF , FMRFamide, myoinhibitory peptide, neurophysin-isotocin, neuropeptide F, and SIFamide precursor that were not found in the transcriptome of the D.

We found transcripts encoding for four of the same neuropeptide receptors, i. In addition, we found transcripts encoding for 10 other neuropeptide receptors, i. Supporting evidence for these gene assignments in the I. Comparing the transcripts from the transcriptomes versus the conspecific genes, we found S1 , S3 , S4 , S15 ; S17 ; S18 ; S20 ; and For the transcript encoding for sulfakinin, the closest match, Although not a neuropeptide, another noteworthy finding was the occurrence of transcripts encoding for pro-protein convertase in the two Illumina transcriptomes 3 in each, respectively similar to that found in D.

Proprotein convertase is essential for the conversion of neuropeptide hormones to the mature form and their subsequent secretions [41] by endoproteolytic cleavage [27]. S13 , calcitonin receptor, S10 , pyrokinin receptor, S5 , Similarly, we only found a transcript encoding for one orcokinin orcokinin 5 , whereas transcripts encoding for 4 different orcokinins were found in the D.

For supporting evidence, see Fig. S6 , eclosion hormone, These are messages for neuropeptides that were not detected in the D. We also found a transcript encoding for neuropeptide F, the importance of which is discussed below; for supporting evidence, see Fig. Also of interest was the finding of the transcript encoding for the cardioacceleratory peptide CCAP receptor see Fig. The finding of the transcript encoding for the CCAP receptor, suggests that the message for the corresponding CCAP peptide may also be present even though not detected in the transcriptomes.

Finally, also noteworthy was the finding of transcripts predicting ion transport peptide Fig. S23 , a molecule important for chloride transport and water reabsorption in insects and, presumably, in many other organisms. Our findings suggest that in ticks, including I. Another possible regulator of tick water balance is inotocin. Although found in many insects [42] , we did not find transcripts encoding for their occurrence in the I. The hormonal regulation of development during blood feeding and reproduction in ticks is not well understood.

Messages encoding several of the neuropeptide hormones or their receptors known to regulate these processes in insects [43] were found in the transcriptomes of the I. Most of these same neuropeptides were reported to be differentially expressed in the D. The presence of messages encoding for allatostatin and allatotropin in the I. However, there is evidence that some elements of the JH pathway occur in ticks Roe, R. Bissinger et al [26] showed that allatostatin was significantly upregulated after mating and feeding to repletion, suggesting a role in tick reproduction.

Also of interest were the messages encoding for eclosion hormone, bursicon and corazonin in adult female I. These hormones are associated with ecdysis and cuticle sclerotization in insects. However, ixodid ticks, including I. Another significant finding was that of a transcript matching the receptor for pyrokinin in I.

It functions in diverse roles in insects [45] but there is no reported evidence of a specific functional role in ticks. The gene for pyrokinin was also reported to occur in the I. Several neuropeptides are known to have broad physiological functions, not specific to any one organ or organ process.

We did not find any evidence of the leucokinin receptor reported to occur in the D. However, ETH expression is not believed to occur in the synganglion. However, immunoreactive staining alone may not be compelling evidence of ETH gene expression. Combining the BLAST matches from the three transcriptomes enabled us to predict a total of 15 expressed neuropeptides. Comparison of the transcripts encoding for neuropeptides receptors found in the three different transcriptomes shows that many more were found by Illumina sequencing, specifically 11 neuropeptides 18 transcripts in sample Il-1 and 9 neuropeptides in sample Il-2 11 transcripts than were found by pyrosequencing, specifically 2 neuropeptides 2 transcripts.

This was also the case for neuropeptide receptors, with 8 predicted 19 transcripts in sample Il-1 , 13 in sample Il-2 25 transcripts versus only 3 4 transcripts in sample , respectively. A message encoding for only one neuropeptide, proctolin, was found solely by pyrosequencing.

Messages encoding for only 2 neuropeptides, allatostatin and allatotropin, and only 2 neuropeptide receptors, calcitonin, and tachykinin were found by both Illumina and pyrosequencing methods. Clearly, although there were benefits obtained by each method of high throughput sequencing, messages encoding for many more neuropeptides and their receptors, with a higher frequency of specific transcripts, were obtained using the Illumina platform than by pyrosequencing.

Transcripts encoding receptors for 6 different types of neurotransmitters were recognized, namely, acetylcholine Ach , gamma aminobutyric acid GABA , dopamine, glutamate, octopamine, and serotonin. Transcripts encoding for acetylcholine muscarinic and nicotinic receptors and 3 types of glutamate receptors had the highest frequency in the I.

Transcripts encoding two different types of GABA receptors ion-channel and metabotropic and three different types of glutamate receptors ionotropic, metabotropic and NMDA were recognized Table 5. In addition, transcripts encoding the enzyme acetylcholinesterase, which degrades acetylcholine, also were recognized. Sequence alignments supporting the functional assignments of these transcripts are shown in Fig. ACh and the enzyme AChE occur in most animals [50] including ticks [14] , [20] , [26] , [51].

Transcripts encoding both muscarinic See Fig. S24 and nicotinic receptors were found. Many more transcripts encoding for these molecules were found in the transcriptome of the I. Bissinger et al. The transcriptomes of the I. Most were found by Illumina sequencing; no transcripts encoding for GABA receptors were found by pyrosequencing Table 5. Transcripts encoding for glutamate receptors were the most abundant Glutamate is a major excitatory neurotransmitter in the nervous system and at the neuromuscular junction of insects and crustaceans.

It can act via inotropic and metabotropic receptors. In insects, muscle contraction is controlled by the glutamatergic pathway [55]. Two glutamate-gated chloride channel receptors were reported in the synganglion of R. In contrast, a remarkable number of transcripts matching sequences for glutamate receptors, including all three types NDMA, ionotropic and metabotropic , were found in the transcriptomes of the I. S27 , Fig. S28 and Fig. S29 for sequence alignments. In addition, numerous transcripts for glutamate synthase were identified 6 in sample Il-1, 3 in sample Il-2 and 1in sample , not shown in Table 5.

It is not clear why so many more glutamate receptor and glutamate synthase transcripts were uncovered by Illumina than Table 5. Transcripts encoding for the neurotransmitter receptors for the monamines dopamine, octopamine and serotonin were also very numerous in the I.

In insects, dopamine functions in modulation of memory recall and motor behavior [56]. In ticks, one of the most important functions of dopamine is stimulation of salivary secretion via the dopaminergic pathway [24] , [54]. Dopamine was also reported to stimulate cuticle plasticization during blood feeding, which is essential for allowing growth in body size [57]. Transcripts encoding for two dopamine receptors were reported in the transcriptome of the D. Here we report 11 transcripts encoding genes for dopamine receptors found in the I.

S25 for sequence alignment. An octopamine receptor was found in the D. Here we report the frequent occurrence of transcripts encoding for octopamine receptors in the synganglion of I. The neurotransmitter serotonin 5-hydroxytryptamine is believed to occur in ticks, based on evidence of serotonin immunoreactivity in the tick nervous system [19]. Only one transcript encoding for a receptor for serotonin was found sample Il-1 Fig.

S31 ; no transcripts encoding for serotonin receptors were found in either of the other two samples. In cockroaches, it is believed that these transporters are involved in the secretion of the NaCl-rich primary saliva [58] , a finding import for ticks in which regulation of the salt composition of saliva is essential for control of body water balance Table 5. In addition to the neuropeptide receptors described above, other GPCRs were identified in the three transcriptomes.

This includes neuropeptide F, found in samples Il-1 and Il-2, as noted previously Table 4. Neuropeptide F has multiple physiological roles, including feeding, metabolism, reproduction and stress responses [46]. This is the first report of its occurrence in the synganglion of ticks. Its role in these parasites remains to be established.

Transcripts encoding GPCRs functioning as pheromone odorant and gustatory receptors were also identified in the I. This is surprising since sensory receptors are part of the peripheral nervous system and normally found in sensory organs near or at the exterior of the body and appendages. However, evidence of atypical expression of such genes, at least the gustatory receptors, has been reported in Drosophila [53]. Since the sex pheromone of I.

The same may be true for the gustatory receptors Table 6. In addition to these 3 GPCRs, transcripts encoding for numerous other GPCRs for which no specific function has been assigned were also found, namely 28 in sample Il-1, 49 in sample Il-2 and 1 in sample A transcript predicting an ecdysone nuclear receptor was identified in the transcriptome of the part-fed female synganglion Il-2 but not in either of the other two transcriptomes.

Alignment of this transcript with the I. However, following mating and rapid engorgement to repletion, its concentrations increase greatly [59]. The much higher expression of this hormone stimulates vitellogenesis and oogenesis [60]. Thus, the expression of this receptor only in sample Il-2 part-fed female synganglion does not appear to be consistent with these reported phenotypic effects.

However, expression of an ecdysone-activated ribosomal protein L63 was found in both Illumina transcriptomes not shown in Table 7 , suggesting that ecdysone activity was prevalent in both the virgin and mated female synganglia. A transcript encoding for an ecdysone receptor was also found in the synganglion of D. Transcripts encoding for other steroid receptors of unknown identity were found in all three transcriptomes while one transcript encoding for juvenile hormone esterase binding protein were found in sample Il-1 and 2 transcripts encoding for this protein were found in sample Il-2 Table 7.

Transcripts for steroid reductase dehydrogenase were also found in all 3 transcriptomes. Transcripts predicting 5 different genes with functions associated specifically with sperm or spermatogenesis were expressed in the female synganglion transcriptomes. Of the 5 different genes in this category, transcripts encoding for epididymal secretory protein E1 similar to Nieman-Pick C2 protein , spermidine synthase see Fig.

S33 , n-acetyl spermine spermidine oxidase and major sperm protein were also found in the D. Transcripts predicting expression of these male-specific genes in the female synganglion of this tick is unexpected. However, examination of the sequence for the spermatogenesis-associated protein revealed that it contained a domain also found in the SSP protein family, commonly found in spermatids and suggesting a function in fertility regulation.

Spermidine was reported to enhance neuronal differentiation in cultures of insect tissues crickets, Acheta domesticus. However, it is not known whether this effect also occurs in vivo [61]. In mosquitoes, e. Epididymal secretory protein E1 is reported to regulate capacitation in mammals [64]. Whether similar phenomena occur in ticks is unknown Table 8. Its expression in blood feeding ticks has also been found to be important as an antimicrobial factor [65] , [66]. Multiple transcripts encoding ferritin found in the I.

PGRPs are members of a broad class of pathogen-associated molecular pattern PAMPs recognition proteins and often initiate signaling activity that leads to inhibition or lysis of the target microbes. Defensins are most often expressed in the hemocytes. However, the I. Thus, the occurrence of the transcript predicting defensin E value 2. Microplusin is a copper chelating molecule that acts as a bacteriostatic rather than a lytic antimicrobial peptide and is known to act against Gram-positive bacteria [68].