alcohol testicles injection what does that do to the testicles

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Alcohol diluent provides the optimal formulation for calcium chloride non-surgical sterilization in dogs

Acta Veterinaria Scandinavica volume 56, Article number:62 (2014) Cite this article

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Abstract

Background

Surgical castration is widely used to sterilize male dogs, just has significant impacts on time to perform the performance, recovery of the animals as well as cost, which tin limit population control programs. Previous enquiry has shown intratesticular injection of calcium chloride dihydrate (CaCl₂) in saline to be a promising alternative to surgery. However, long-term azoospermia was not maintained at dosages depression enough to avoid side furnishings. In the search for an optimized formulation, the electric current investigation is the get-go study on long-term sterilization effects of intratesticular injection of CaCl₂ in either lidocaine solution or alcohol in dogs. CaCl₂ at 20% concentration in lidocaine solution or alcohol was administered via intratesticular injection to groups of 21 dogs each. The treated animals were examined at 2, 6, and 12 months for sperm production, claret levels of testosterone, and side furnishings; at time zero and 12 months for testicular size and semen volume. The experimentally treated animals were compared to a control grouping receiving saline injection only.

Results

Testicles of dogs treated with CaCl_ii in either diluent significantly decreased in size. After assistants of CaCl₂ in lidocaine solution, sterility was achieved for at least 12 months in 75% of treated dogs. However, optimal long-term contraceptive effectiveness was achieved with CaCl₂ in booze, which resulted in azoospermia over the 12-calendar month written report period. Testosterone levels significantly decreased following treatment with CaCl_two, and sexual practice disappeared. Although testosterone returned to baseline levels past 12 months for the group treated with CaCl₂ in lidocaine, dogs injected with CaCl_two in alcohol had a 63.6% drib in testosterone level, which remained at the low finish of physiological range throughout the study. No adverse furnishings were noted.

Conclusions

A single, bilateral intratesticular injection of xx% CaCl₂ in 95% ethanol was a reliable method for induction of sterilization in 18-28 kg male dogs in this report. The approach showed long-term efficacy and reduced sexual beliefs. This chemical method of sterilization might provide an effective, efficient culling to surgical castration that tin can take positive impacts on dog welfare.

Background

Canine overpopulation remains a trouble facing many countries throughout the world. Culling methods to surgical sterilization that are effective, piece of cake to administer, prophylactic, and affordable would offer immense benefits, allowing animate being welfare organizations, public health programs, and governments to reach farther with limited resources [1].

An intratesticular injection of calcium chloride dihydrate (CaCl_two) in solution represents a promising method for non-surgical sterilization [2]-[7]. A previous dose-determination study reported that a xx% solution of CaCl₂ in saline demonstrated good long-term efficacy without the undesirable side effects that occurred with college dosages [2]. These findings partially confirmed the results of short-term, histology-based studies on CaCl₂ past other investigators who used a xx% concentration [3],[5]-[7]. All the same, when 20% CaCl₂ in saline solution, as typically used for sterilization, was evaluated for efficacy over a longer period, the effect was not permanent: sperm product returned in some of the treated dogs, and testosterone levels increased to baseline levels by 12 months following injection [2]. For CaCl₂ to be used finer for canine sterilization, the conception must exist optimized to ensure permanent azoospermia.

The effectiveness of CaCl₂ every bit a sterilizing agent may be augmented by the diluent used in the formulation. The primeval published abstract on the use of intratesticular injection of CaCl_ii for sterilization in a diversity of animals reported that aqueous solutions permitted higher concentrations, but tinctures in 80%-99% alcohol had the advantages of less pain, less peripheral inflammation, and more consistent results [8]. Intratesticular assistants of a tincture of CaCl_two in ethanol in dogs was reported to have anesthetic properties, in comparing with saline solutions of CaCl₂[4]. Also, booze is known to have contained utility every bit a sterilant. In 1998, Yoon and Yoon [9] institute that chemic castration with alcohol alone was as constructive as orchiectomy in reducing testosterone levels in blood of rats. Furthermore, a single injection of 95% ethanol directly into the vas deferens caused atrophy of the cauda epididymis. All-encompassing necrosis and exfoliation of the seminiferous elements were conspicuous [x], with irreversible obstructive necrosis [eleven],[12].

In addition, solutions of CaCl₂ in other diluents have been used. Samanta and Jana reported on the effectiveness of lidocaine derivatives equally diluents for CaCl₂ chemosterilization in dogs and cats [5]-[7]. For instance, using ane% lignocaine hydrochloride equally a base for intratesticular injections of CaCl₂ in dogs resulted in complete degeneration of germ cells in a 45-day trial [half-dozen]. The investigators reported that these changes may take been due to the necrotizing backdrop of CaCl₂ and/or the significant reduction in intratesticular and claret levels of testosterone.

Despite the promising results on the use of CaCl_two as a nonsurgical sterilization method, picayune is known near long-term effectiveness or impact on canis familiaris wellness and behavior. This lack of information has hampered the widespread awarding of CaCl_two to accost the trouble of dog overpopulation.

The objective of the current study was to evaluate the long-term (i.e., i year) efficacy of intratesticular injection of 20% CaCl_two in booze versus lidocaine for the relative ability to halt sperm production and reduce blood levels of testosterone in dogs. Nosotros hypothesized a greater effectiveness for 1 or both formulations, as compared to historical utilize of 20% CaCl₂ in saline solitary.

Methods

Animals

For the study, 52 healthy, owned, mixed-breed male person dogs living in a shelter were selected. The dogs were 2 to 6 years of age (mean = 3.5 years, SD = ane.1 years) and weighed xviii to 28 kg (hateful = 22.9 kg, SD = 2.93 kg). Good health condition was confirmed by routine claret testing and clinical exam. To assess the fertility of the dogs, an andrological examination (including physical and ultrasonographic examination and evaluation of semen quality) was performed before the showtime of the written report. Every dog showed sexual interest when exposed to a bitch in estrus.

Dogs were routinely de-wormed and vaccinated. The dogs were housed in private shelters, fed standard commercial dog nutrient twice per day, given water ad libitum, and not subjected to changes in habits during the report. Dogs were housed in groups of iii in a comfortable chief enclosure with outdoor runs. Indoor infinite had temperature maintained above 15°C and below 26°C and relative humidity ranging from 30% to 70%.

Investigations were conducted in accordance with the Principles for the Intendance and Use of Inquiry Animals, promulgated past the Eu. The Italian Ministry of Health (Progetto di Ricerca corrente 2009 IZS SI 11/09: "Randagismo applicazione e valutazione di metodi innovativi per il controllo delle nascite") approved this written report.

Experimental protocol

At day 0 (T₀), the animals were randomly assigned to iii groups using a random number tabular array: 2 experimental groups (A and B) of 21 dogs each and a control group (C) of x dogs. The get-go author was aware of group assignment, only technicians collecting data on the subjects were bullheaded to status. Semen evaluation and collection of blood samples were performed. Subsequently, dogs were lightly sedated with an intramuscular (IM) injection of 5-10 mg of acepromazine maleate (Prequillan, Fatro, Italy) per 10 kg of torso weight. The testicular widths were measured with a caliper. According to the scrotal width, the right dosage of solution was injected into each testicle (see "Preparation and intratesticular injection of CaCl₂ solution"). Dogs in group A were injected with CaCl_ii in a solution containing 1% lidocaine chlorhydrate. Dogs in group B were injected with CaCl_ii in alcohol. Dogs in group C were injected with a saline solution.

At 2, 6, and 12 months (T_i, T₂, T₃, respectively), semen evaluation was performed, and blood samples were taken for testosterone evaluation. At 12 months (T_three), testicular width was measured. Throughout the trial, the dogs were under clinical observation.

Preparation and intratesticular injection of CaCl2 solution

To prepare the solution containing 20% CaCl_ii and one% lidocaine, 20 g of CaCl₂ dihydrate powder (Sigma Aldrich Corporation) was added to a last volume of 100 mL with a i% solution of lidocaine chlorhydrate (Salp spa, Italian republic), mixed, and sterilized in Falcon tubes.

The alcohol solution of twenty% CaCl2 dihydrate was prepared equally follows: 20 g of CaCl2 dihydrate powder (Sigma Aldrich Corporation) was brought to a final volume of 100 mL of 95% ethanol (Bakery Analyzed ACS, JT Baker), mixed, and sterilized in Falcon tubes.

The dogs received a single, bilateral intratesticular injection of solution (Effigy 1) proportional to testicular width: animals with scrotal diameters of 19-22 millimeters (mm) wide received 0.viii mL injections, whereas animals with scrotal diameters of at least 23 mm wide received 1 mL injections [2].

Figure 1

Intratesticular injection. Photo shows procedure of unmarried, bilateral intratesticular injection of 1 mL of 20% CaCl₂ in ethanol for sterilization of mature male person dogs. Each injection was performed using a sterile 22-estimate needle that was directed from the ventral aspect of each testis approximately 0.5 cm from the epididymal tail towards the cranial aspect of that testis. The solution was carefully deposited along the entire route by linear infiltration, while withdrawing needle from proximal to distal finish.

Full size epitome

Semen volume, total sperm count and motility

Semen was collected by digital manipulation of the penis using plastic cones (artificial vaginas) (IMV Technologies, Italian republic) into sterile graduated tubes at 37°C [ii],[xiii],[14]. Ejaculate volume was measured to include all 3 semen fractions obtained. Inside 30-60 min semen was examined by computer-assisted sperm analysis (CASA) (IVOS Version 12.2; Hamilton Thorne Biosciences Inc., Beverly, MA, USA), which was validated for a large range of sperm counts [15],[16]. Total sperm count and movement were obtained. Results were confirmed past optical microscopy evaluation.

Assay for serum testosterone

To determine testosterone levels at time intervals T₀ to T₃, dogs received subcutaneous (SC) injections of i,000 international units (I.U.) of human chorionic gonadotropin (hCG) (Creative Biomart, CD, Inc.) [2],[17]. At 120 min after the hCG injections, blood was collected every bit previously described [ii]. Testosterone was measured by a chemiluminescence technique (Immulite Immunoassay System, Siemens).

Routine clinical observations

All the animals were kept under routine clinical observations from T₀ to T₃. After the chemical sterilization process, continuous observations were conducted for the first 72 hours, followed by daily observations for up to 15 days, followed by observations as indicated by the study protocol. The parameters evaluated during clinical observation included physiological data (respiratory rate, salivation, torso weight, appetite, rectal temperature, etc.), response to palpation, posture, vox, mental status (submissive, etc.). Behaviors indicative of pain or discomfort, sexual beliefs (mounting) and aggressive behavior (growling, snapping) were carefully evaluated [xviii].

Measurement of testicular width

Scrotal width was used as an index of testicular size [19]. At T₀ and T₃, widths (mm) of the correct and left testes were measured using laboratory calipers. Data were expressed as a mean betwixt the width of left and right testicles.

Statistical analyses

All data were summarized for each private canine field of study by measurement (weight, testosterone level, semen volume, total sperm count, sperm motion, testicular width), grouping (A, B, C), and time point (T₀, T₁, T₂, T₃) using the Microsoft Excel 2011 program (Microsoft Corporation, Redmond, Washington, The states). The boilerplate of the testicular width measurements were used for analysis. These data were described in terms of the boilerplate and standard deviation (SD) and presented as mean ± SD in the results for brevity.

Statistical analyses were conducted using Statistica (StatSoft, Inc. Tulsa, OK, Us). Repeated measures of analysis of variance (ANOVA), with Fourth dimension as the inside factor and Group as the between factor, were used to evaluate the measurements in the three groups (A-C) beyond iv time points (T₀, T_ane, T_ii, T₃) for testosterone, full sperm count and move, or two time points (T₀ and T₃) for semen volume and testicular width. If the result of the overall test showed significance, then planned comparisons were conducted. Dunnett's test for comparison to a control group was used, besides every bit univariate or multivariate planned comparisons to determine if the measures inverse after treatment and if the treated groups differed from the command grouping. A two-tailed significance level of P < 0.05 was identified.

Results

Routine clinical observation

Before the injection of sterilant or control saline, the hateful weight of the dogs was 22.viii ± 2.9kg. No changes in body weight during the trial were observed. For all dogs, values for hematology and clinical chemistry consistently remained within reference ranges.

All animals in the study tolerated the intratesticular injections of CaCl_ii. Hurting parameters did not differ during the written report for nearly dogs. A few dogs, however, showed signs of small-scale pain at needle puncture of the scrotum during the injection: 2% of dogs injected with either CaCl_two in alcohol, lidocaine solution, or normal saline had abdominal muscle contraction, and 1% vocalized. The minor transient discomfort probably was caused past needle puncture of the scrotum or fluid pressure over the testicular capsule. Scrotal ultrasonography (USG) revealed a hypoechoic intratesticular surface area, corresponding to a collection of the injected fluid (Effigy 2).

Effigy 2

Scrotal ultrasonography subsequently intratesticular injection of CaCl _two . A hypoechoic intratesticular area corresponding to a collection of the injected fluid was observed.

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Even if the injection was performed advisedly, seepage occurred in a few dogs. However, the solution was wiped away immediately with dry out gauze, and no agin effects were noticed after the seepage.

During the commencement 2 weeks after the CaCl₂ injection, the dogs in groups A and B and the control dogs (grouping C) did not experience any agitation, fever, or marked inflammatory swelling of the testis or changes in evaluated parameters. No agin side effects were noticed at the 2-week period. Notwithstanding, get-go after 24 hours following injection and continuing for the first 3-4 days, a slight increase in firmness of testes on palpation was noticed in dogs in groups A and B and the control dogs (group C); the increased firmness was slightly more noticeable in dogs in group A. From ane week to approximately 1.5 months in dogs in groups A and B, atrophy of the testes gradually progressed, leaving a small fibrotic remnant. An interference with sexual behavior (i.east., loss of libido, mounting and dominance behavior) and aggression was observed in groups A and B following treatment. In contrast, no testicular changes or alteration of beliefs were observed in the control group, C.

Total sperm count, sperm move and semen volume

At T_0, the mean full sperm count (x10^half-dozen) was 346.2 ± 33.nine in group A, 348.4 ± 32.iv in grouping B, and 335.9 ± 34.9 in the control group. Analysis of variance procedures indicated a pregnant interaction of Grouping and Time for full sperm count (F = 276; P < 0.001). Farther analyses revealed that this result was due to reduced total sperm count for experimentally treated dogs, but not for the control dogs. No significant variation in full sperm count was noticed at T₁, T_ii, and T_three in the control group that had received saline injection (F = 1.eight; P = 0.18) (340 ± 23.3; 313.5 ± 40.v; 311.4 ± 21.iv, respectively).

Although full sperm count in the experimental groups A and B did not differ from that of the control group C at baseline T₀ (F = 0.94; P = 0.338), both experimental groups had significantly lower total sperm count than did the controls at T_i-T₃ (F = 11476; P < 0.001).

In groups A and B, all dogs were azoospermic at T₁ and T₂. At T₃, 17 (81%) dogs in grouping A were azoospermic, and 4 dogs (nineteen%) were severely oligospermic (40.5 ± 5.1), exhibiting merely 5% movement (Table 1). The mean total sperm count in dogs of group A at T₃ was vii.7 ± 16.four. At T_three, all dogs of group B were azoospermic. According to statistical analysis, a meaning reduction was observed in total sperm counts later on intratesticular injection of CaCl₂ to dogs in group A (F = 2220; p < 0.001) and B (F = 2283; P < 0.009) (Table 1).

Table 1 Furnishings of intratesticular injection of calcium chloride on reproductive parameters at i year post-injection

Full size tabular array

At T₀, move was ninety% in group A, 95% in group B, and 80% in the command grouping (C). Sperm movement in the control group was lxxx% at all times tested (T₁-T_iii). Of the 42 treated dogs, only four that were injected with CaCl₂ in lidocaine had 5% sperm motility (Tabular array 1). Statistical analysis was not possible due to a lack of variability in the information.

Ejaculate volume was not significantly different beyond groups at T₀ (Group A: two.98 ± 0.55; Group B: 3.15 ± 0.68; Grouping C: 3.45 ± 0.51). Assay revealed a meaning Time by Group interaction (F = 46.2, P < 0.001) in which the groups treated with CaCl₂ had lower semen volume at T₃ than the control group C (F = 63.5, P < 0.001) (Group A: two.36 ± 0.53; Group B: ane.64 ± 0.55; Grouping C: 3.50 ± 0.51). Semen volumes in both group A (F = 39.5, P < 0.001) and group B (F = 239.9, P < 0.001) were significantly reduced from T₀ to T₃.

Assay of serum testosterone

At T₀, the hateful values of testosterone levels (ng/dL) were 456.2 ± 132.4 in grouping A, 454.6 ± 159.ix in group B, and 721.2 ± 176.two in the command group (C). For all dogs tested, testosterone values remained within physiological range (100-k ng/dL) [17] throughout the course of the study, although a single intratesticular injection of CaCl₂ was sufficient to decrease plasma testosterone concentrations significantly in the treated dogs. Analyses revealed an overall issue of Grouping by Fourth dimension (F = 10.9; P < 0.001), with treated groups having lower testosterone after injection than did the control group (F = 165.7; P < 0.001). In dissimilarity, changes in the serum testosterone levels in the control group were not statistically meaning (P > 0.05).

Figure iii depicts the levels of serum testosterone graphically over fourth dimension. Following the injection of CaCl₂ in lidocaine solution (group A), testosterone decreased significantly for vi months (F = 0.47; P < 0.003), although levels at T₃ returned to baseline. At 12 months following injection, testosterone levels for the group treated with CaCl₂ in alcohol (Group B) dropped 63.6%, as compared to baseline. Testosterone levels in group B decreased significantly and remained at the low cease of the physiological range throughout the 12-month follow-up period (F = 65.1; P < 0.001).

Effigy iii

Furnishings of intratesticular injection of CaCl ₂ on serum testosterone levels over time. Following the injection of CaCl₂ in lidocaine solution (group A), testosterone decreased significantly (F = 0.47; P < 0.003) for up to six months, although testosterone levels at 12 months returned to baseline. Later on injection of calcium chloride in alcohol (grouping B), testosterone levels decreased significantly (F = 65.1, P < 0.001) throughout the 12-month follow-up flow.

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Measurement of testicular width

Testicular width also varied by Group and Time (F = 412; P < 0.001). Average testicular width at baseline (T₀) was similar across groups. The control group showed no meaning divergence in testicular width over fourth dimension (P > 0.05).

At T₀ versus (vs) T₃, the mean values of testicular width (mm) were 24.7 ± ane.5 vs 12.8 ± 1.0 in grouping A, 24.8 ± i.5 vs 12.2 ± 0.9 in group B, and 24.9 ± two.1 vs 24.9 ± 2.i in the control group (C) (Figure four).

Figure 4

Changes in testicular width afterward intratesticular injection of CaCl ₂ . At 12 months (T₃) later handling with CaCl₂ (group A and group B), significant reductions in testicular width were observed (*P < 0.001), as compared with no or minimal changes seen in the command (C) group.

Full size image

After treatment, the width of scrota had declined significantly in both of the CaCl_ii-treated groups (A: F = 2036; P < 0.001 and B: F = 2235; P < 0.001) and was narrower than that of the command group (F = 802; P < 0.001). The boilerplate reduction in testicular width at T_three was approximately 50% in both group A and group B.

Word

The aim of the current research was to study any potential improvement in the efficacy of using CaCl₂ as a nonsurgical sterilization method due to the chemical nature of the solvents. In this report, ii diluents (lidocaine or alcohol) were tested for use with CaCl₂ every bit a sterilant for dogs. Our results indicate that alcohol was a superior solution for CaCl₂ assistants, resulting in complete azoospermia over a 12-calendar month menses, decreased sexual behavior, and no side effects.

Alcohol lone is a chemical that causes testicular sclerosis. A study of intratesticular injection of accented alcohol in rats demonstrated that levels of testosterone were as low as in surgically castrated rats [9]. Studies of ethanol solutions of CaCl₂ have demonstrated the definite advantages of more consistent efficacy, less pain, and less peripheral inflammation [viii].

In our study involving canine male contraception, no or minimal signs of discomfort were observed post-obit injection, with variation dependent on the agent injected. Minor transient pain occurred during the injection, as any needle inserted through skin will crusade somatic hurting for an instant. The explanation for the relative lack of discomfort following the injection is that afferent nerve endings associated with pain sensation are located on the scrotal peel and in the sheathing of the testis, rather than within the testicular and epididymal parenchyma [20]. Given the beefcake of the testes, astringent testicular pain when experienced is visceral and triggered past rapid pressure deforming the testicular capsule. During chemical castration, it is important to deliver the injection very slowly to avert triggering the testicular pressure receptors. In our experience, dogs that had been injected with the alcohol tincture of CaCl_two exhibited less discomfort on the day following the injection than those injected with the lidocaine diluent.

Sperm analysis revealed that the injection of CaCl₂ in booze had long-term effectiveness at one year post-treatment, whereas the injection of CaCl_two in lidocaine solution was effective in all dogs for vi months. At the i-twelvemonth time point, some of the dogs that had been treated with CaCl_ii in lidocaine solution regained residual production of sperm. All the same, we cannot affirm that these dogs regained fertility, because the severe oligospermia and poor motility of sperm were unlikely to outcome in impregnation. Nevertheless it is not possible to exclude that these dogs might regain sufficient sperm product in the future. Regeneration of seminiferous tubules has been reported eight weeks after handling with v% concentrations of CaCl₂, but non at higher dosages [5]. However, our long-term study found that sperm was produced in at least some dogs injected with CaCl_ii in lidocaine. Thus, our findings differ from reports of brusk-term studies of like concentrations of CaCl₂ that concluded that `permanent' sterilization had occurred [5],[half dozen].

Semen volume decreased significantly in dogs injected with CaCl₂. This was due to a reduction of the second sperm rich fraction of the ejaculate and indicates poor semen quality.

For contraception of stray male dogs, desirable methods crave a sufficient reduction in the level of testosterone and, therefore, suppression of sexual behavior. Although previous enquiry on CaCl_ii in both diluents demonstrated a statistically significant decrease in serum testosterone [five]-[7], it was not stated explicitly whether the testosterone levels had decreased to below that of physiological range. Prior investigations on the use of CaCl₂ in lidocaine solution reported the necrotizing backdrop of CaCl₂, resulting in low serum concentrations of testosterone [6],[7].

In the electric current written report and our previous piece of work [ii], a significant decrease in testosterone in all CaCl₂-treated groups was measured, despite the fact that serum testosterone remained inside normal physiological levels over a 12 month period. In the current study, we also observed the disappearance of ambitious and sex-related behavior in the treated dogs throughout the written report. To our knowledge, the modify in level of testosterone needed to consequence in a pregnant decrease in or absence of canine sexual behavior has never been quantified. From the current written report, a reduction in testosterone levels to the low finish of the physiological range was sufficient to affect behavior. This is of import because a reduction in aggression and sexual behavior is unremarkably sought in canine sterilization programs.

The electric current study is the commencement to evaluate the long-term furnishings of unlike diluents used in CaCl₂ sterilization. Our findings demonstrate the high potential of 20% CaCl_ii in booze as a sterilant for use in stray male dogs. The sterilant fulfills the master requirements for application to a population of stray canines. A single, bilateral intratesticular injection for stray dogs is effective in achieving long-term infertility, inhibits sexual behavior, does not cause chronic stress to the creature, causes few inflammatory reactions, lacks other undesirable side furnishings, is hands performed, and is economical.

Conclusions

A unmarried, bilateral intratesticular injection of 20% CaCl₂ in booze produced azoospermia in all dogs at one year, representing an optimal method for sterilization in male dogs, whereas the effects of CaCl₂ in lidocaine solution lasted for only six months. The sterilization arroyo using CaCl₂ in alcohol resulted in a durable reduction of testosterone, as compared to baseline levels, and reduced ambitious and sexual behavior. Intratesticular injection of CaCl₂ in booze appears to be an effective and reliable sterilization method in male dogs, making it a proficient potential alternative to surgical castration. Nevertheless more studies on a larger and more variable population of dogs in a wider weight range as well equally in roaming dogs are needed to ameliorate understand the applicability of this sterilization method on devious dogs.

Abbreviations

ANOVA:

Analysis of variance

CaCl₂:

Calcium chloride

X g:

Centrifugal strength X gravity

°C:

Centigrade

dL:

Deciliter

F:

F-statistic (Fisher)

G:

Gauge

g:

Gram

hCG:

Human chorionic gonadotrophin

I.U.:

International unit of measurement

IM:

Intramuscular

kg:

Kilogram

x:

Magnification

μL:

Microliter

mg:

Milligram

mm:

Millimeter

p:

p-value

SD:

Standard deviation

SEM:

Standard fault of the hateful

SC:

Subcutaneous

vs:

Versus

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Acknowledgements

The authors are deeply grateful to Parsemus Foundation, Berkeley, California, USA for fiscal help, continuous support, and interest in this report. Likewise, the authors acknowledge Linda Brent for analysis and interpretation of data and language review and Holly Abrams for editing the paper.

Author information

Affiliations

1. Department of Emergency and Organ Transplantation (DETO), Section of Veterinary Clinic and Animal Product, University of Bari Aldo Moro, SP per Casamassima km 3, Valenzano, BA, Italian republic

   Raffaella Leoci, Giulio Aiudi, Fabio Silvestre & Giovanni One thousand Lacalandra

2. Parsemus Foundation, Berkeley, 94702, CA, The states

   Elaine A Lissner

Corresponding writer

Correspondence to Raffaella Leoci.

Additional information

Competing interests

The authors declare that they take no competing interests.

Authors' contributions

RL was involved in the concept and design of the report, analysis and estimation of results, semen sampling and evaluation, and preparation of this manuscript. RL, GA, EAL were involved in the revision of study design. RL and GA performed the intratesticular injection of the dogs. GA and FS performed the ultrasonography. FS was involved in the clinical intendance of the dogs, blood sampling, and acquisition of data. GML, EAL, LR were involved in revision of the manuscript. All authors have read and approved the manuscript.

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Leoci, R., Aiudi, G., Silvestre, F. et al. Alcohol diluent provides the optimal formulation for calcium chloride non-surgical sterilization in dogs. Acta Vet Scand 56, 62 (2014). https://doi.org/10.1186/s13028-014-0062-2

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• Received: 06 June 2013

• Accepted: 05 September 2014

• Published: 14 October 2014

• DOI : https://doi.org/ten.1186/s13028-014-0062-2

Keywords

• Calcium chloride
• Canine
• Chemical castration
• Dog
• Nonsurgical sterilization
• Population management

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